Phages produce persisters.

Arguably, the greatest threat to bacteria is phages. It is often assumed that those bacteria that escape phage infection have mutated or utilized phage-defence systems; however, another possibility is that a subpopulation forms the dormant persister state in a manner similar to that demonstrated for bacterial cells undergoing nutritive, oxidative, and antibiotic stress. Persister cells do not undergo mutation and survive lethal conditions by ceasing growth transiently.

CRISPR-Cas Controls Cryptic Prophages.

The bacterial archetypal adaptive immune system, CRISPR-Cas, is thought to be repressed in the best-studied bacterium, K-12. We show here that the CRISPR-Cas system is active and serves to inhibit its nine defective (i.e.

Xenogeneic silencing relies on temperature-dependent phosphorylation of the host H-NS protein in Shewanella.

Lateral gene transfer (LGT) plays a key role in shaping the genome evolution and environmental adaptation of bacteria. Xenogeneic silencing is crucial to ensure the safe acquisition of LGT genes into host pre-existing regulatory networks. We previously found that the host nucleoid structuring protein (H-NS) silences prophage CP4So at warm temperatures yet enables this prophage to excise at cold temperatures in Shewanella oneidensis.

Persister Cells Resuscitate Using Membrane Sensors that Activate Chemotaxis, Lower cAMP Levels, and Revive Ribosomes.

Persistence, the stress-tolerant state, is arguably the most vital phenotype since nearly all cells experience nutrient stress, which causes a sub-population to become dormant. However, how persister cells wake to reconstitute infections is not understood well. Here, using single-cell observations, we determined that Escherichia coli persister cells resuscitate primarily when presented with specific carbon sources, rather than spontaneously.

Cyanide bioremediation: the potential of engineered nitrilases.

The cyanide-degrading nitrilases are of notable interest for their potential to remediate cyanide contaminated waste streams, especially as generated in the gold mining, pharmaceutical, and electroplating industries. This review provides a brief overview of cyanide remediation in general but with a particular focus on the cyanide-degrading nitrilases. These are of special interest as the hydrolysis reaction does not require secondary substrates or cofactors, making these enzymes particularly good candidates for industrial remediation processes.

Residue Y70 of the Nitrilase Cyanide Dihydratase from Is Critical for Formation and Activity of the Spiral Oligomer.

Nitrilases pose attractive alternatives to the chemical hydrolysis of nitrile compounds. The activity of bacterial nitrilases towards substrate is intimately tied to the formation of large spiral-shaped oligomers. In the nitrilase CynD (cyanide dihydratase) from , mutations in a predicted oligomeric surface region altered its oligomerization and reduced its activity.

Bacillus pumilus Cyanide Dihydratase Mutants with Higher Catalytic Activity.

Cyanide degrading nitrilases are noted for their potential to detoxify industrial wastewater contaminated with cyanide. However, such application would benefit from an improvement to characteristics such as their catalytic activity and stability. Following error-prone PCR for random mutagenesis, several cyanide dihydratase mutants from Bacillus pumilus were isolated based on improved catalysis.

Probing an Interfacial Surface in the Cyanide Dihydratase from Bacillus pumilus, A Spiral Forming Nitrilase.

Nitrilases are of significant interest both due to their potential for industrial production of valuable products as well as degradation of hazardous nitrile-containing wastes. All known functional members of the nitrilase superfamily have an underlying dimer structure. The true nitrilases expand upon this basic dimer and form large spiral or helical homo-oligomers.

Orphan toxin OrtT (YdcX) of Escherichia coli reduces growth during the stringent response.

Toxin/antitoxin (TA) systems are nearly universal in prokaryotes; toxins are paired with antitoxins which inactivate them until the toxins are utilized. Here we explore whether toxins may function alone; i.e.

Probing C-terminal interactions of the Pseudomonas stutzeri cyanide-degrading CynD protein.

The cyanide dihydratases from Bacillus pumilus and Pseudomonas stutzeri share high amino acid sequence similarity throughout except for their highly divergent C-termini. However, deletion or exchange of the C-termini had different effects upon each enzyme. Here we extended previous studies and investigated how the C-terminus affects the activity and stability of three nitrilases, the cyanide dihydratases from B.

The MqsR/MqsA toxin/antitoxin system protects Escherichia coli during bile acid stress.

Toxin/antitoxin (TA) systems are ubiquitous within bacterial genomes, and the mechanisms of many TA systems are well characterized. As such, several roles for TA systems have been proposed, such as phage inhibition, gene regulation and persister cell formation. However, the significance of these roles is nebulous due to the subtle influence from individual TA systems.

Phosphodiesterase DosP increases persistence by reducing cAMP which reduces the signal indole.

Persisters are bacteria that are highly tolerant to antibiotics due to their dormant state and are of clinical significance owing to their role in infections. Given that the population of persisters increases in biofilms and that cyclic diguanylate (c-di-GMP) is an intracellular signal that increases biofilm formation, we sought to determine whether c-di-GMP has a role in bacterial persistence. By examining the effect of 30 genes from Escherichia coli, including diguanylate cyclases that synthesize c-di-GMP and phosphodiesterases that breakdown c-di-GMP, we determined that DosP (direct oxygen sensing phosphodiesterase) increases persistence by over a thousand fold.

Toxin YafQ increases persister cell formation by reducing indole signalling.

Persister cells survive antibiotic and other environmental stresses by slowing metabolism. Since toxins of toxin/antitoxin (TA) systems have been postulated to be responsible for persister cell formation, we investigated the influence of toxin YafQ of the YafQ/DinJ Escherichia coli TA system on persister cell formation. Under stress, YafQ alters metabolism by cleaving transcripts with in-frame 5'-AAA-G/A-3' sites.

Draft Genome Sequence of Cupriavidus sp. Strain SK-3, a 4-Chlorobiphenyl- and 4-Clorobenzoic Acid-Degrading Bacterium.

We report the draft genome sequence of Cupriavidus sp. strain SK-3, which can use 4-chlorobiphenyl and 4-clorobenzoic acid as the sole carbon source for growth. The draft genome sequence allowed the study of the polychlorinated biphenyl degradation mechanism and the recharacterization of the strain SK-3 as a Cupriavidus species.

Draft Genome Sequence of Cupriavidus sp. Strain SK-4, a di-ortho-Substituted Biphenyl-Utilizing Bacterium Isolated from Polychlorinated Biphenyl-Contaminated Sludge.

Cupriavidus sp. strain SK-4 is a bacterium capable of growing aerobically on monochlorobiphenyls and dichlorobiphenyls as the sole carbon sources for growth. Here, we report its draft genome sequence with the aim of facilitating an understanding of polychlorinated biphenyl biodegradation mechanisms.

RalR (a DNase) and RalA (a small RNA) form a type I toxin-antitoxin system in Escherichia coli.

For toxin/antitoxin (TA) systems, no toxin has been identified that functions by cleaving DNA. Here, we demonstrate that RalR and RalA of the cryptic prophage rac form a type I TA pair in which the antitoxin RNA is a trans-encoded small RNA with 16 nucleotides of complementarity to the toxin mRNA. We suggest the newly discovered antitoxin gene be named ralA for RalR antitoxin.

Toxin GhoT of the GhoT/GhoS toxin/antitoxin system damages the cell membrane to reduce adenosine triphosphate and to reduce growth under stress.

Toxin/antitoxin (TA) systems perhaps enable cells to reduce their metabolism to weather environmental challenges although there is little evidence to support this hypothesis. Escherichia coli GhoT/GhoS is a TA system in which toxin GhoT expression is reduced by cleavage of its messenger RNA (mRNA) by antitoxin GhoS, and TA system MqsR/MqsA controls GhoT/GhoS through differential mRNA decay. However, the physiological role of GhoT has not been determined.

Interactions of the TnaC nascent peptide with rRNA in the exit tunnel enable the ribosome to respond to free tryptophan.

A transcriptional attenuation mechanism regulates expression of the bacterial tnaCAB operon. This mechanism requires ribosomal arrest induced by the regulatory nascent TnaC peptide in response to free L-tryptophan (L-Trp). In this study we demonstrate, using genetic and biochemical analyses, that in Escherichia coli, TnaC residue I19 and 23S rRNA nucleotide A2058 are essential for the ribosome's ability to sense free L-Trp.

Catabolic plasmid specifying polychlorinated biphenyl degradation in Cupriavidus sp. strain SK-4: mobilization and expression in a pseudomonad.

Strain SK-4, a polychlorinated biphenyl (PCB) degrader previously reported to utilize di-ortho-substituted biphenyl, was genotypically re-characterized as a species of Cupriavidus. The bacterium harbored a single plasmid (pSK4), which resisted curing and which, after genetic marking by a transposon (SK4Tn5), could be mobilized into a pseudomonad. Analysis of pSK4 in both the transconjugant and the wild type revealed that it specifies the genes coding for 2-hydroxy-2,4-pentadienoate degradation in addition to those of the upper biphenyl pathway.

Arrested protein synthesis increases persister-like cell formation.

Biofilms are associated with a wide variety of bacterial infections and pose a serious problem in clinical medicine due to their inherent resilience to antibiotic treatment. Within biofilms, persister cells comprise a small bacterial subpopulation that exhibits multidrug tolerance to antibiotics without undergoing genetic change. The low frequency of persister cell formation makes it difficult to isolate and study persisters, and bacterial persistence is often attributed to a quiescent metabolic state induced by toxins that are regulated through toxin-antitoxin systems.

Type II toxin/antitoxin MqsR/MqsA controls type V toxin/antitoxin GhoT/GhoS.

Toxin endoribonucleases of toxin/antitoxin (TA) systems regulate protein production by selectively degrading mRNAs but have never been shown to control other TA systems. Here we demonstrate that toxin MqsR of the MqsR/MqsA system enriches toxin ghoT mRNA in vivo and in vitro, since this transcript lacks the primary MqsR cleavage site 5'-GCU. GhoT is a membrane toxin that causes the ghost cell phenotype, and is part of a type V TA system with antitoxin GhoS that cleaves specifically ghoT mRNA.

Draft genome sequence of the cyanide-utilizing bacterium Pseudomonas fluorescens strain NCIMB 11764.

We report here the 6.97-Mb draft genome sequence of Pseudomonas fluorescens strain NCIMB 11764, which is capable of growth on cyanide as the sole nitrogen source. The draft genome sequence allowed the discovery of several genes implicated in enzymatic cyanide turnover and provided additional information contributing to a better understanding of this organism's unique cyanotrophic ability.