Regulatory mechanisms involved in the aggregation of pseudomonas aeruginosa cells

Abstract

The vast majority of microbial growth occurs in multicellular consortia known as biofilms. Biofilms have often been defined as adherent to an inert or living surface, although suspended aggregates were recently posited as biofilms based on their resistance to antibiotics and the host immune response. The association of suspended aggregates of Pseudomonas aeruginosa cells with chronic human infections (e.g. chronic wounds and the cystic fibrosis lung) is generating increased interest in this phenotype. However, the majority of biofilm studies have focused on surface attached biofilms, and therefore this study was undertaken to investigate the molecular mechanisms involved in aggregation. It was previously demonstrated that exposure to the surfactant sodium dodecyl-sulfate (SDS) could induce aggregation of P. aeruginosa cells. SDS may be used as a proxy for bacterial stress due to its ability to damage membranes and proteins, a situation which may be encountered in diverse environments including human infectious disease. It has been shown that a novel SiaAD pathway and the nucleotide based second messenger cyclic di-guanosine monophosphate (c-di-GMP) are important determinants of aggregation. The work presented here has characterised the proteins SiaB and SiaC as regulators of SDS induced aggregation. The results suggest that a complex signal transduction cascade, operating primarily through posttranslational activation of SiaABCD, leads to the output of elevated c-di-GMP levels and cellular aggregation. Effectors of the SiaABCD regulatory pathway have previously been identified, including the CupA, Psl and CdrA cell surface adhesins that promote aggregate formation. Activation of the SiaABCD pathway appears to regulate the expression of these effectors at the posttranscriptional level. This regulation was found to involve a global posttranscriptional regulator in the mRNA binding protein RsmA. The interconnectivity between SiaABCD and RsmA mediated regulation is complex, and includes feedback regulation of SiaABCD to achieve homeostasis. Finally, the SiaABCD pathway was shown to regulate aggregation in the absence of SDS, such as during growth with glucose or during exposure to the lung surfactant phosphatidylcholine. The loss of siaACD was subsequently demonstrated to reduce the survival of nematodes (Caenorhabditis elegans) feeding on the respective mutant strains in a slow killing model of infection.