A highly conductive fibre network enables centimetre-scale electron transport in multicellular cable bacteria.

Biological electron transport is classically thought to occur over nanometre distances, yet recent studies suggest that electrical currents can run along centimetre-long cable bacteria. The phenomenon remains elusive, however, as currents have not been directly measured, nor have the conductive structures been identified. Here we demonstrate that cable bacteria conduct electrons over centimetre distances via highly conductive fibres embedded in the cell envelope. Direct electrode measurements reveal nanoampere currents in intact filaments up to 10.1 mm long (>2000 adjacent cells). A network of parallel periplasmic fibres displays a high conductivity (up to 79 S cm-1), explaining currents measured through intact filaments. Conductance rapidly declines upon exposure to air, but remains stable under vacuum, demonstrating that charge transfer is electronic rather than ionic. Our finding of a biological structure that efficiently guides electrical currents over long distances greatly expands the paradigm of biological charge transport and could enable new bio-electronic applications.

Targeting mechanisms of tailed bacteriophages.

Phages differ substantially in the bacterial hosts that they infect. Their host range is determined by the specific structures that they use to target bacterial cells. Tailed phages use a broad range of receptor-binding proteins, such as tail fibres, tail spikes and the central tail spike, to target their cognate bacterial cell surface receptors. Recent technical advances and new structure-function insights have begun to unravel the molecular mechanisms and temporal dynamics that govern these interactions. Here, we review the current understanding of the targeting machinery and mechanisms of tailed phages. These new insights and approaches pave the way for the application of phages in medicine and biotechnology and enable deeper understanding of their ecology and evolution.

Can disc diffusion susceptibility tests assess the antimicrobial activity of engineered nanoparticles?

The use of disc diffusion susceptibility tests to determine the antibacterial activity of engineered nanoparticles (ENPs) is questionable because their low diffusivity practically prevents them from penetrating through the culture media. In this study, we investigate the ability of such a test, namely the Kirby-Bauer disc diffusion test, to determine the antimicrobial activity of Au and Ag ENPs having diameters from 10 to 40 nm on Escherichia coli cultures. As anticipated, the tests did not show any antibacterial effects of Au nanoparticles (NPs) as a result of their negligible diffusivity through the culture media. Ag NPs on the other hand exhibited a strong antimicrobial activity that was independent of their size. Considering that Ag, in contrast to Au, dissolves upon oxidation and dilution in aqueous solutions, the apparent antibacterial behavior of Ag NPs is attributed to the ions they release. The Kirby-Bauer method, and other similar tests, can therefore be employed to probe the antimicrobial activity of ENPs related to their ability to release ions rather than to their unique size-dependent properties. Graphical abstractᅟ.

Bistability in a metabolic network underpins the de novo evolution of colony switching in Pseudomonas fluorescens.

Phenotype switching is commonly observed in nature. This prevalence has allowed the elucidation of a number of underlying molecular mechanisms. However, little is known about how phenotypic switches arise and function in their early evolutionary stages. The first opportunity to provide empirical insight was delivered by an experiment in which populations of the bacterium Pseudomonas fluorescens SBW25 evolved, de novo, the ability to switch between two colony phenotypes. Here we unravel the molecular mechanism behind colony switching, revealing how a single nucleotide change in a gene enmeshed in central metabolism (carB) generates such a striking phenotype. We show that colony switching is underpinned by ON/OFF expression of capsules consisting of a colanic acid-like polymer. We use molecular genetics, biochemical analyses, and experimental evolution to establish that capsule switching results from perturbation of the pyrimidine biosynthetic pathway. Of central importance is a bifurcation point at which uracil triphosphate is partitioned towards either nucleotide metabolism or polymer production. This bifurcation marks a cell-fate decision point whereby cells with relatively high pyrimidine levels favour nucleotide metabolism (capsule OFF), while cells with lower pyrimidine levels divert resources towards polymer biosynthesis (capsule ON). This decision point is present and functional in the wild-type strain. Finally, we present a simple mathematical model demonstrating that the molecular components of the decision point are capable of producing switching. Despite its simple mutational cause, the connection between genotype and phenotype is complex and multidimensional, offering a rare glimpse of how noise in regulatory networks can provide opportunity for evolution.

Founder niche constrains evolutionary adaptive radiation.

Adaptive radiation of a lineage into a range of organisms with different niches underpins the evolution of life's diversity. Although the role of the environment in shaping adaptive radiation is well established, theory predicts that the evolvability and niche of the founding ancestor are also of importance. Direct demonstration of a causal link requires resolving the independent effects of these additional factors. Here, we accomplish this using experimental bacterial populations and demonstrate how the dynamics of adaptive radiation are constrained by the niche of the founder. We manipulated the propensity of the founder to undergo adaptive radiation and resolved the underlying causal changes in both its evolvability and niche. Evolvability did not change, but the propensity for adaptive radiation was altered by changes in the position and breadth of the niche of the founder. These observations provide direct empirical evidence for a link between the niche of organisms and their propensity for adaptive radiation. This general mechanism may have rendered the evolutionary dynamics of extant adaptive radiations dependent on chance events that determined their founding ancestors.

The evolutionary emergence of stochastic phenotype switching in bacteria.

Stochastic phenotype switching - or bet hedging - is a pervasive feature of living systems and common in bacteria that experience fluctuating (unpredictable) environmental conditions. Under such conditions, the capacity to generate variable offspring spreads the risk of being maladapted in the present environment, against offspring likely to have some chance of survival in the future. While a rich subject for theoretical studies, little is known about the selective causes responsible for the evolutionary emergence of stochastic phenotype switching. Here we review recent work - both theoretical and experimental - that sheds light on ecological factors that favour switching types over non-switching types. Of particular relevance is an experiment that provided evidence for an adaptive origin of stochastic phenotype switching by subjecting bacterial populations to a selective regime that mimicked essential features of the host immune response. Central to the emergence of switching types was frequent imposition of 'exclusion rules' and 'population bottlenecks' - two complementary faces of frequency dependent selection. While features of the immune response, exclusion rules and bottlenecks are likely to operate in many natural environments. Together these factors define a set of selective conditions relevant to the evolution of stochastic switching, including antigenic variation and bacterial persistence.

The distribution of fitness effects of new beneficial mutations in Pseudomonas fluorescens.

Theoretical studies of adaptation emphasize the importance of understanding the distribution of fitness effects (DFE) of new mutations. We report the isolation of 100 adaptive mutants-without the biasing influence of natural selection-from an ancestral genotype whose fitness in the niche occupied by the derived type is extremely low. The fitness of each derived genotype was determined relative to a single reference type and the fitness effects found to conform to a normal distribution. When fitness was measured in a different environment, the rank order changed, but not the shape of the distribution. We argue that, even with detailed knowledge of the genetic architecture underpinning the adaptive types (as is the case here), the DFEs remain unpredictable, and we discuss the possibility that general explanations for the shape of the DFE might not be possible in the absence of organism-specific biological details.

Experimental evolution of bet hedging.

Bet hedging-stochastic switching between phenotypic states-is a canonical example of an evolutionary adaptation that facilitates persistence in the face of fluctuating environmental conditions. Although bet hedging is found in organisms ranging from bacteria to humans, direct evidence for an adaptive origin of this behaviour is lacking. Here we report the de novo evolution of bet hedging in experimental bacterial populations. Bacteria were subjected to an environment that continually favoured new phenotypic states. Initially, our regime drove the successive evolution of novel phenotypes by mutation and selection; however, in two (of 12) replicates this trend was broken by the evolution of bet-hedging genotypes that persisted because of rapid stochastic phenotype switching. Genome re-sequencing of one of these switching types revealed nine mutations that distinguished it from the ancestor. The final mutation was both necessary and sufficient for rapid phenotype switching; nonetheless, the evolution of bet hedging was contingent upon earlier mutations that altered the relative fitness effect of the final mutation. These findings capture the adaptive evolution of bet hedging in the simplest of organisms, and suggest that risk-spreading strategies may have been among the earliest evolutionary solutions to life in fluctuating environments.

Immigration history controls diversification in experimental adaptive radiation.

Diversity in biological communities is a historical product of immigration, diversification and extinction, but the combined effect of these processes is poorly understood. Here we show that the order and timing of immigration controls the extent of diversification. When an ancestral bacterial genotype was introduced into a spatially structured habitat, it rapidly diversified into multiple niche-specialist types. However, diversification was suppressed when a niche-specialist type was introduced before, or shortly after, introduction of the ancestral genotype. In contrast, little suppression occurred when the same niche specialist was introduced relatively late. The negative impact of early arriving immigrants was attributable to the historically sensitive outcome of interactions involving neutral competition and indirect facilitation. Ultimately, the entire boom-and-bust dynamics of adaptive radiation were altered. These results demonstrate that immigration and diversification are tightly linked processes, with small differences in immigration history greatly affecting the evolutionary emergence of diversity.

Novel nirK cluster genes in Nitrosomonas europaea are required for NirK-dependent tolerance to nitrite.

Nitrite reductase (NirK) of Nitrosomonas europaea confers tolerance to nitrite (NO2-). The nirK gene is clustered with three genes of unknown physiological function: ncgABC. At present, this organization is unique to nitrifying bacteria. Here we report that the ncgABC gene products facilitate NirK-dependent NO2- tolerance by reversing the negative physiological effect that is associated with the activity of NirK in their absence. We hypothesize that the ncg gene products are involved in the detoxification of nitric oxide that is produced by NirK.

Expression of nitrite reductase in Nitrosomonas europaea involves NsrR, a novel nitrite-sensitive transcription repressor.

Production of nitric oxide (NO) and nitrous oxide (N(2)O) by ammonia (NH(3))-oxidizing bacteria in natural and man-made habitats is thought to contribute to the undesirable emission of NO and N(2)O into the earth's atmosphere. The NH(3)-oxidizing bacterium Nitrosomonas europaea expresses nitrite reductase (NirK), an enzyme that has so far been studied predominantly in heterotrophic denitrifying bacteria where it is involved in the production of these nitrogenous gases. The finding of nirK homologues in other NH(3)-oxidizing bacteria suggests that NirK is widespread among this group; however, its role in these nitrifying bacteria remains unresolved. We identified a gene, nsrR, which encodes a novel nitrite (NO(2) (-))-sensitive transcription repressor that plays a pivotal role in the regulation of NirK expression in N. europaea. NsrR is a member of the Rrf2 family of putative transcription regulators. NirK was expressed aerobically in response to increasing concentrations of NO(2) (-) and decreasing pH. Disruption of nsrR resulted in the constitutive expression of NirK. NsrR repressed transcription from the nirK gene cluster promoter (P(nir)), the activity of which correlated with NirK expression. Reconstruction of the NsrR-P(nir) system in Escherichia coli revealed that repression by NsrR was reversed by NO(2) (-) in a pH-dependent manner. The findings are consistent with the hypothesis that N. europaea expresses NirK as a defence against the toxic NO(2) (-) that is produced during nitrification.

Nitrosomonas europaea expresses a nitric oxide reductase during nitrification.

In this paper, we report the identification of a norCBQD gene cluster that encodes a functional nitric oxide reductase (Nor) in Nitrosomonas europaea. Disruption of the norB gene resulted in a strongly diminished nitric oxide (NO) consumption by cells and membrane protein fractions, which was restored by the introduction of an intact norCBQD gene cluster in trans. NorB-deficient cells produced amounts of nitrous oxide (N2O) equal to that of wild-type cells. NorCB-dependent activity was present during aerobic growth and was not affected by the inactivation of the putative fnr gene. The findings demonstrate the presence of an alternative site of N2O production in N. europaea.

Nitrite reductase of Nitrosomonas europaea is not essential for production of gaseous nitrogen oxides and confers tolerance to nitrite.

A gene that encodes a periplasmic copper-type nitrite reductase (NirK) was identified in Nitrosomonas europaea. Disruption of this gene resulted in the disappearance of Nir activity in cell extracts. The nitrite tolerance of NirK-deficient cells was lower than that of wild-type cells. Unexpectedly, NirK-deficient cells still produced nitric oxide (NO) and nitrous oxide (N(2)O), the latter in greater amounts than that of wild-type cells. This demonstrates that NirK is not essential for the production of NO and N(2)O by N. europaea. Inactivation of the putative fnr gene showed that Fnr is not essential for the expression of nirK.