Biology subjects

Granato, E. T.

Publications and source records attributed to Granato, E. T..

6 recordsLinked to original sources

bioRxiv2025

Type VI secretion system activity at lethal antibiotic concentrations leads to overestimation of weapon potency

Competition assays are a mainstay of modern microbiology, offering a simple and cost-effective means to quantify microbe-microbe interactions in vitro. Here we demonstrate a key weakness of this method that arises when competing microbes interact via toxins, such as those secreted via the type VI secretion system (T6SS). Timelapse microscopy reveals T6SS-armed Acinteobacter baylyi bacteria can maintain lethal T6SS activity against E. coli target cells, even under selective conditions intended to eliminate A. baylyi. Further, this residual killing creates a density- and T6SS-dependent bias in the apparent recovery of E. coli, leading to a misreporting of competition outcomes especially where target survival is low. We also show that incubating A. baylyi / E. coli co-cultures in liquid antibiotic prior to selective plating can substantially correct this bias. Our findings demonstrate the need for caution when using selective plating as part of T6SS competition assays, or assays involving other toxin-producing bacteria.

microbiology↗

bioRxiv2024

Horizontal gene transfer can reshape bacterial warfare

Bacteria commonly use molecular weaponry to kill or inhibit competitors. Genes encoding many of these weapons can be transmitted horizontally, but the impact on bacterial warfare is not understood. Here, we develop an ecological model of bacterial warfare where there is potential for horizontal gene transfer. Our model predicts that weapon gene transfer from an attacker to a target strain is possible, but at a low rate such that transfer has a negligible impact on competition outcomes. We tested the model empirically using a transmissible plasmid encoding colicin E2, a potent antibacterial toxin produced by Escherichia coli. As predicted by the model, we find that toxin plasmid transfer is feasible during warfare, but the resulting transconjugants remain rare. However, exploring the model further reveals realistic conditions where transfer has major impacts. Specifically, the model predicts that whenever competing strains have access to some unique nutrients, transconjugants can proliferate and reach high abundances. In support of these predictions, short- and long-term experiments show that transconjugants can thrive when nutrient competition is relaxed. Our work shows how horizontal gene transfer can reshape bacterial warfare in a way that can benefit a weapon gene but not the bacteria that carries it.

microbiology↗

bioRxiv2022

Collective protection against the type VI secretion system in bacteria

Bacteria commonly face attacks from other strains using the type VI secretion system (T6SS), a molecular speargun that stabs and intoxicates competitors. Here we show how bacteria can work together to collectively defend themselves against these attacks. This project began with an outreach activity: while developing an online computer game of bacterial warfare, we noticed that one strategist ( Slimy) that made extracellular polymeric substances (EPS) was able to resist attacks from another strategist that employed the T6SS ( Stabby). This observation motivated us to model this scenario more formally, using dedicated agent-based simulations. The model predicts that EPS production can serve as a collective defence mechanism, which protects both producing cells and neighbouring cells that do not make EPS. We then tested our model with a synthetic community that contains a T6SS-wielding attacker (Acinetobacter baylyi), and two T6SS-sensitive target strains (Escherichia coli) that either secrete EPS, or not. As predicted by our modelling, we find that the production of EPS leads to collective protection against T6SS attacks, where EPS producers protect each other and nearby non-producers. We identify two processes that explain this protection: EPS sharing between cells and a second general mechanism whereby groups of resistant cells shield susceptible cells ( flank protection). Our work shows how EPS-producing bacteria can work together to defend themselves from the type VI secretion system.

microbiology↗

bioRxiv2020

The Evolution of Mass Cell Suicide in Bacterial Warfare

Behaviours that reliably cause the death of an actor are typically strongly disfavoured by natural selection, and yet many bacteria undergo cell lysis to release anti-competitor toxins [1-4]. This behaviour is most easily explained if only a few cells die to release toxins and help their clonemates, but the number of cells that actually lyse during bacterial warfare is unknown. The challenge is that one cannot distinguish cells that have undergone programmed suicide from those that were simply killed by a competitors toxin. We developed a two-colour fluorescence reporter assay in Escherichia coli to overcome this problem. Surprisingly, this revealed conditions where nearly all cells undergo programmed lysis. Adding a DNA-damaging toxin (DNase colicin) to a focal strain causes it to engage in mass cell suicide where around 85% of cells lyse to release their own toxin. Time-lapse 3D confocal microscopy revealed that self-lysis occurs at even higher frequencies (~94%) at the interface between competing colonies. We sought to understand how such high levels of cell suicide could be favoured by natural selection. Exposing E. coli that do not perform lysis to the DNase colicin revealed that mass lysis only occurs when cells are going to die anyway from toxin exposure. From an evolutionary perspective, this renders the behaviour cost-free as these cells have zero reproductive potential. This explains how mass cell suicide can evolve, as any small benefit to surviving clonemates can lead to the strategy being favoured by natural selection. Our findings have strong parallels to the suicidal attacks of social insects [5-8], which are also performed by individuals with low reproductive potential, suggesting convergent evolution in these very different organisms. HIGHLIGHTSO_LIA novel assay can detect Escherichia coli undergoing cell suicide to release toxins C_LIO_LIWe quantified the frequency of suicidal self-lysis during competitions C_LIO_LIUnder some conditions, nearly all cells will self-lyse to release toxins C_LIO_LISelf-lysis makes evolutionary sense as cells will die anyway from competitors toxins C_LI

microbiology↗

bioRxiv2018

Virulence evolution in the opportunistic bacterial pathogen Pseudomonas aeruginosa

Bacterial opportunistic pathogens are feared for their difficult-to-treat nosocomial infections and for causing morbidity in immunocompromised patients. Here, we study how such a versatile opportunist, Pseudomonas aeruginosa, adapts to conditions inside and outside its model host Caenorhabditis elegans, and use phenotypic and genotypic screens to identify the mechanistic basis of virulence evolution. We found that virulence significantly dropped in unstructured environments both in the presence and absence of the host, but remained unchanged in spatially structured environments. Reduction of virulence was either driven by a substantial decline in the production of siderophores (in treatments without hosts) or toxins and proteases (in treatments with hosts). Whole-genome sequencing of evolved clones revealed positive selection and parallel evolution across replicates, and showed an accumulation of mutations in regulator genes controlling virulence factor expression. Our study identifies the spatial structure of the non-host environment as a key driver of virulence evolution in an opportunistic pathogen.

microbiology↗

bioRxiv2017

The path to re-evolve cooperation is constrained in Pseudomonas aeruginosa

BackgroundA common form of cooperation in bacteria is based on the secretion of beneficial metabolites, shareable as public good among cells at the group level. Because cooperation can be exploited by \"cheat\" mutants, which contribute less or nothing to the public good, there has been great interest in understanding the conditions required for cooperation to remain evolutionarily stable. In contrast, much less is known about whether cheats, once fixed in the population, are able to revert back to cooperation when conditions change. Here, we tackle this question by subjecting experimentally evolved cheats of Pseudomonas aeruginosa, partly deficient for the production of the iron-scavenging public good pyoverdine, to conditions previously shown to favor cooperation.\n\nResultsFollowing approximately 200 generations of experimental evolution, we screened 720 evolved clones for changes in their pyoverdine production levels. We found no evidence for the re-evolution of full cooperation, even in environments with increased spatial structure, and reduced costs of cooperation - two conditions that have previously been shown to maintain cooperation. In contrast, we observed selection for complete abolishment of pyoverdine production. The patterns of complete trait degradation were likely driven by \"cheating on cheats\" in unstructured, iron-limited environments where pyoverdine is important for growth, and selection against a maladaptive trait in iron-rich environments where pyoverdine is superfluous.\n\nConclusionsOur study shows that the path to re-evolve cooperation seems constrained. One reason might be that the number of mutational targets potentially leading to reversion is limited. Alternatively, it could be that the selective conditions required for revertants to spread from rare are much more stringent than those needed to maintain cooperation.

evolutionary biology↗