TY - JOUR

T1 - Genetic analysis of functions involved in the late stages of biofilm development in Burkholderia cepacia H111

AU - Huber, Birgit

AU - Riedel, Kathrin

AU - Köthe, Manuela

AU - Givskov, Michael

AU - Molin, Søren

AU - Eberl, Leo

N1 - Keywords: 4-Butyrolactone; Bacterial Proteins; Biofilms; Burkholderia Infections; Burkholderia cepacia; Cystic Fibrosis; DNA Transposable Elements; Gene Expression Regulation, Bacterial; Humans; Hydrophobicity; Image Processing, Computer-Assisted; Microscopy, Confocal; Mutagenesis, Insertional; Polystyrenes; Signal Transduction; Surface Properties

PY - 2002

Y1 - 2002

N2 - Burkholderia cepacia and Pseudomonas aeruginosa often co-exist as mixed biofilms in the lungs of patients suffering from cystic fibrosis (CF). Here, we report the isolation of 13 random mini-Tn5 insertion mutants of B. cepacia H111 that are defective in biofilm formation on a polystyrene surface. We show that the screening procedure used in this study is biased towards mutants defective in the late stages of biofilm development. A detailed quantitative analysis of the biofilm structures formed by wild-type and mutant strains revealed that the isolated mutants are impaired in their abilities to develop a typical three-dimensional biofilm structure. Molecular investigations showed that the genes required for biofilm maturation fall into several classes: (i). genes encoding for surface proteins; (ii). genes involved in the biogenesis and maintenance of an integral outer membrane; and (iii). genes encoding regulatory factors. It is shown that three of the regulatory mutants produce greatly reduced amounts of N-octanoylhomoserine lactone (C8-HSL). This compound serves as the major signal molecule of the cep quorum-sensing system. As this density-dependent regulatory system is involved in the regulation of biofilm maturation, we investigated the interplay between the three regulatory genes and the quorum-sensing cascade. The results of these investigations show that the identified genes encode for regulatory elements that are positioned upstream of the cep system, indicating that the quorum-sensing system of B. cepacia is a major checkpoint for biofilm formation.

AB - Burkholderia cepacia and Pseudomonas aeruginosa often co-exist as mixed biofilms in the lungs of patients suffering from cystic fibrosis (CF). Here, we report the isolation of 13 random mini-Tn5 insertion mutants of B. cepacia H111 that are defective in biofilm formation on a polystyrene surface. We show that the screening procedure used in this study is biased towards mutants defective in the late stages of biofilm development. A detailed quantitative analysis of the biofilm structures formed by wild-type and mutant strains revealed that the isolated mutants are impaired in their abilities to develop a typical three-dimensional biofilm structure. Molecular investigations showed that the genes required for biofilm maturation fall into several classes: (i). genes encoding for surface proteins; (ii). genes involved in the biogenesis and maintenance of an integral outer membrane; and (iii). genes encoding regulatory factors. It is shown that three of the regulatory mutants produce greatly reduced amounts of N-octanoylhomoserine lactone (C8-HSL). This compound serves as the major signal molecule of the cep quorum-sensing system. As this density-dependent regulatory system is involved in the regulation of biofilm maturation, we investigated the interplay between the three regulatory genes and the quorum-sensing cascade. The results of these investigations show that the identified genes encode for regulatory elements that are positioned upstream of the cep system, indicating that the quorum-sensing system of B. cepacia is a major checkpoint for biofilm formation.

M3 - Journal article

C2 - 12406218

VL - 46

SP - 411

EP - 426

JO - Molecular Microbiology

JF - Molecular Microbiology

SN - 0950-382X

IS - 2

ER -