No abstract

Exotoxin A (ETA) is secreted by Pseudomonas aeruginosa under iron-limiting growth conditions. The ETA structural gene, toxA, is regulated at the transcriptional level by the gene products of the regAB operon. The expression of both toxA and regAB is repressed under iron-replete conditions, suggesting a role for the ferric uptake regulator (Fur) in regulation of ETA synthesis; however, the Fur protein does not interact directly with the toxA or the regAB promoters. Evidence is presented that the iron control of ETA synthesis is mediated by a Fur-regulated alternative sigma factor, PvdS, which had initially been identified as a positive activator for the production of the siderophore pyoverdin. In a delta pvdS deletion mutant, ETA was produced at low levels of less than 5% compared to wild type, but still in response to iron starvation, and introduction of a functional pvdS gene on a plasmid fully restored wild-type levels and normal iron regulation of ETA synthesis. Therefore, a functional pvdS locus is essential for ETA production. Neither toxA nor regAB mRNA was detectable in a delta pvdS mutant. Overexpression of pvdS from the tac promoter on a plasmid resulted in a high-level and iron-independent production of ETA in wild-type PAO1, in the delta pvdS strain, but not in a delta regA strain as a host. These findings suggest that PvdS is required for the activation of the regAB promoters. The transcription of regAB and toxA after induction of the P tac-pvdS gene was monitored in cells grown in high-iron medium. While both regAB and toxA were highly expressed during all growth phases under microaerobic conditions, toxA transcripts were detected only during the exponential but not the early stationary phase of growth under aerobic conditions. These results suggest that a second regulatory mechanism besides the Fur-PvdS system is involved in iron regulation of ETA production.

The rapid increase in antibiotic-resistant bacteria has emphasized the urgent need to identify new treatments for bacterial infections. One attractive approach, reducing the need for expensive and time-consuming clinical trials, is to repurpose existing clinically approved compounds for use as antibacterial agents. Nucleoside analogues are commonly used for treating viral and fungal infections, as well as for treating cancers, but have received relatively little attention as treatments for bacterial infections. However, a significant number of clinically approved derivatives of both pyrimidines and purines including halogenated, thiolated, and azolated compounds have been shown to have antibacterial activity. In the small number of studies carried out to date, such compounds have shown promise in treating bacterial infections. Here, we review the mechanisms of action and antibacterial activities of nucleoside analogues that can potentially be repurposed for treating infections as well as considering possible limitations in their usage.

SummaryThe symbiosis island ICEMlSym R7A of Mesorhizobium loti R7A is an integrative and conjugative element (ICE) that carries genes required for a nitrogen-fixing symbiosis with Lotus species. ICEMlSym R7A encodes homologues (TraR, TraI1 and TraI2) of proteins that regulate plasmid transfer by quorum sensing in rhizobia and agrobacteria. Introduction of traR cloned on a plasmid induced excision of ICEMlSym R7A in all cells, a 1000-fold increase in the production of 3-oxo-C6-homoserine lactone (3-oxo-C6-HSL) and a 40-fold increase in conjugative transfer. These effects were dependent on traI1 but not traI2. Induction of expression from the traI1 and traI2 promoters required the presence of plasmid-borne traR and either traI1 or 100 pM 3-oxo-C6-HSL, suggesting that traR expression or TraR activity is repressed in wild-type cells by a mechanism that can be overcome by additional copies of traR. The traI2 gene formed an operon with hypothetical genes msi172 and msi171 that were essential for ICEMlSym R7A excision and transfer. Our data suggest that derepressed TraR in conjunction with TraI1-synthesized 3-oxo-C6-HSL regulates excision and transfer of ICEMlSym R7A through expression of msi172 and msi171. Homologues of msi172 and msi171 were present on putative ICEs in several a-proteobacteria, indicating a conserved role in ICE excision and transfer.

To characterise Pseudomonas aeruginosa populations during chronic lung infections of non-cystic fibrosis bronchiectasis patients, we used whole-genome sequencing to 1) assess the diversity of P. aeruginosa and the prevalence of multilineage infections; 2) seek evidence for cross-infection or common source acquisition; and 3) characterise P. aeruginosa adaptations.189 isolates, obtained from the sputa of 91 patients attending 16 adult bronchiectasis centres in the UK, were whole-genome sequenced.Bronchiectasis isolates were representative of the wider P. aeruginosa population. Of 24 patients from whom multiple isolates were examined, there were seven examples of multilineage infections, probably arising from multiple infection events. The number of nucleotide variants between genomes of isolates from different patients was in some cases similar to the variations observed between isolates from individual patients, implying the possible occurrence of cross-infection or common source acquisition.Our data indicate that during infections of bronchiectasis patients, P. aeruginosa populations adapt by accumulating loss-of-function mutations, leading to changes in phenotypes including different modes of iron acquisition and variations in biofilm-associated polysaccharides. The within-population diversification suggests that larger scale longitudinal surveillance studies will be required to capture cross-infection or common source acquisition events at an early stage.