Investigation of the algT operon sequence in mucoid and non-mucoid Pseudomonas aeruginosa isolates from 115 Scandinavian patients with cystic fibrosis and in 88 in vitro non-mucoid revertants

Pseudomonas aeruginosa is the dominant pathogen causing chronic lung infections in patients with cystic fibrosis (CF). After an initial phase characterized by intermittent colonizations, a chronic infection is established upon conversion of P. aeruginosa from the non-mucoid to the mucoid, alginate-overproducing phenotype. During the chronic infection the isolation of both mucoid and non-mucoid isolates in CIF sputum samples is very common. The purpose of the present study was to establish, by sequence analysis, the types of mutations present in the algTmucABD operon in a large number of mucoid and non-mucoid P. aeruginosa isolates from Scandinavian CIF patients and in in vitro-derived non-mucoid revertants. Mucoid (83) and non-mucoid isolates (103) from 91 Scandinavian patients with chronic P. aeruginosa infection and 24 non-mucoid isolates from intermittently colonized CIF patients were investigated. In addition, 88 spontaneous non-mucoid revertants obtained in vitro from nine mucoid CIF isolates were also included in the study. Mutations in mucA were found in 92% of the mucoid and in up to 70% of the non-mucoid isolates from chronically infected patients, indicating that the majority of non-mucoid isolates are revertants. None of the non-mucoid isolates from intermittently colonized CIF patients harboured mucA mutations. Although algT has been considered an important gene for secondary-site mutations responsible for reversion to non-mucoidy, only 30% of the mucA-mutated non-mucoid CIF isolates had mutations in algT. In contrast, 83% of the in vitro-derived spontaneous non-mucoid revertants had mutations in algT, showing that in the CF lung there is a selection for non-mucoid revertants with secondary-site mutations in genes other than algT In addition, we report, to our knowledge for the first time, loss-of-function mutations in the negative regulators mucB and mucD in CIF clinical isolates. In some of the CIF isolates these mutations are associated with moderate alginate production. In conclusion, most non-mucoid isolates from chronically infected CIF patients are revertants and the mechanism of revertance is algT-independent in the CIF lung.