Positive Signature-Tagged Mutagenesis in Pseudomonas aeruginosa: Tracking Patho-Adaptive Mutations Promoting Airways Chronic Infection

PLoS Pathogens

Roger C. Levesque¹  Irene Bianconi²  Moira Paroni²  Cristina Cigana²  Alessandra Bragonzi²  Giovanni Bertoni³  Andrea Milani³ 

[1] Institut de biologie intégrative et des systèmes (IBIS), Faculté de médecine et Pavillon Charles-Eugène Marchand, Université Laval, Québec, Canada;San Raffaele Scientific Institute, Division of Immunology, Transplantation and Infectious Diseases, Milano, Italy;Università degli Studi di Milano, Department of Biomolecular Sciences and Biotechnologies, Milano, Italy

关键词: Pseudomonas aeruginosa;    Respiratory infections;    Animal models of infection;    Mouse models;    Phenotypes;    Secretion;    Bacterial biofilms;    Pathogen motility;   

DOI  :  10.1371/journal.ppat.1001270

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

The opportunistic pathogen Pseudomonas aeruginosa can establish life-long chronic infections in the airways of cystic fibrosis (CF) patients. Persistent lifestyle is established with P. aeruginosa patho-adaptive variants, which are clonal with the initially-acquired strains. Several reports indicated that P. aeruginosa adapts by loss-of-function mutations which enhance fitness in CF airways and sustain its clonal expansion during chronic infection. To validate this model of P. aeruginosa adaptation to CF airways and to identify novel genes involved in this microevolution, we designed a novel approach of positive-selection screening by PCR-based signature-tagged mutagenesis (Pos-STM) in a murine model of chronic airways infection. A systematic positive-selection scheme using sequential rounds of in vivo screenings for bacterial maintenance, as opposed to elimination, generated a list of genes whose inactivation increased the colonization and persistence in chronic airways infection. The phenotypes associated to these Pos-STM mutations reflect alterations in diverse aspects of P. aeruginosa biology which include lack of swimming and twitching motility, lack of production of the virulence factors such as pyocyanin, biofilm formation, and metabolic functions. In addition, Pos-STM mutants showed altered invasion and stimulation of immune response when tested in human respiratory epithelial cells, indicating that P. aeruginosa is prone to revise the interaction with its host during persistent lifestyle. Finally, sequence analysis of Pos-STM genes in longitudinally P. aeruginosa isolates from CF patients identified signs of patho-adaptive mutations within the genome. This novel Pos-STM approach identified bacterial functions that can have important clinical implications for the persistent lifestyle and disease progression of the airway chronic infection.

【 授权许可】

CC BY