Identification of 17 Pseudomonas aeruginosa sRNAs and prediction of sRNA-encoding genes in 10 diverse pathogens using the bioinformatic tool sRNAPredict2

sRNAs are small, non-coding RNA species that control numerous cellular processes. Although it iswidely accepted that sRNAs are encoded by most if not all bacteria, genome-wide annotations for sRNA-encoding genes have been conducted in only a few of the nearly 300 bacterial species sequenced to date. To facilitate the efficient annotation of bacterial genomes for sRNA-encoding genes, we developed a program, sRNAPredict2, that identifies putative sRNAs by searching for co-localization of genetic features commonly associated with sRNA-encoding genes. Using sRNAPredict2, we conducted genome-wide annotations for putative sRNA-encoding genes in the intergenic regions of 11 diverse pathogens. In total, 2759 previously unannotated candidate sRNA loci were predicted. There was considerable range in the number of sRNAs predicted in the different pathogens analyzed, raising the possibility that there are species-specific differences in the reliance on sRNA-mediated regulation. Of 34 previously unannotated sRNAs predicted in the opportunistic pathogen Pseudomonas aeruginosa, 31 were experimentally tested and 17 were found to encode sRNA transcripts. Our findings suggest that numerous genes have been missed in the current annotations of bacterial genomes and that, by using improved bioinformatic approaches and tools, much remains to be discovered in ‘intergenic’ sequences

Detection of and Response to Signals Involved in Host-Microbe Interactions by Plant-Associated Bacteria

Diverse interactions between hosts and microbes are initiated by the detection of host-released chemical signals. Detection of these signals leads to altered patterns of gene expression that culminate in specific and adaptive changes in bacterial physiology that are required for these associations. This concept was first demonstrated for the members of the family Rhizobiaceae and was later found to apply to many other plant-associated bacteria as well as to microbes that colonize human and animal hosts. The family Rhizobiaceae includes various genera of rhizobia as well as species of Agrobacterium. Rhizobia are symbionts of legumes, which fix nitrogen within root nodules, while Agrobacterium tumefaciens is a pathogen that causes crown gall tumors on a wide variety of plants. The plant-released signals that are recognized by these bacteria are low-molecular-weight, diffusible molecules and are detected by the bacteria through specific receptor proteins. Similar phenomena are observed with other plant pathogens, including Pseudomonas syringae, Ralstonia solanacearum, and Erwinia spp., although here the signals and signal receptors are not as well defined. In some cases, nutritional conditions such as iron limitation or the lack of nitrogen sources seem to provide a significant cue. While much has been learned about the process of host detection over the past 20 years, our knowledge is far from being complete. The complex nature of the plant-microbe interactions makes it extremely challenging to gain a comprehensive picture of host detection in natural environments, and thus many signals and signal recognition systems remain to be described

Analysis of Secretin-Induced Stress in Pseudomonas aeruginosa Suggests Prevention Rather than Response and Identifies a Novel Protein Involved in Secretin Function ▿

Secretins are bacterial outer membrane proteins that are important for protein export. However, they can also mislocalize and cause stress to the bacterial cell, which is dealt with by the well-conserved phage shock protein (Psp) system in a highly specific manner. Nevertheless, some bacteria have secretins but no Psp system. A notable example is Pseudomonas aeruginosa, a prolific protein secretor with the potential to produce seven different secretins. We were interested in investigating how P. aeruginosa might deal with the potential for secretin-induced stress without a Psp system. Microarray analysis revealed the absence of any transcriptional response to XcpQ secretin overproduction. However, transposon insertions in either rpoN, truB, PA4068, PA4069, or PA0943 rendered P. aeruginosa hypersensitive to XcpQ production. The PA0943 gene was studied further and found to encode a soluble periplasmic protein important for XcpQ localization to the outer membrane. Consistent with this, a PA0943 null mutation reduced the levels of type 2 secretion-dependent proteins in the culture supernatant. Therefore, this work has identified a novel protein required for normal secretin function in P. aeruginosa. Taken together, all of our data suggest that P. aeruginosa lacks a functional equivalent of the Psp stress response system. Rather, null mutations in genes such as PA0943 may cause increased secretin-induced stress to which P. aeruginosa cannot respond. Providing the PA0943 mutant with the ability to respond, in the form of critical Psp proteins from another species, alleviated its secretin sensitivity