Parallel compensatory evolution stabilizes plasmids across the parasitism-mutualism continuum

Plasmids drive genomic diversity in bacteria via horizontal gene transfer [1 and 2]; nevertheless, explaining their survival in bacterial populations is challenging [3]. Theory predicts that irrespective of their net fitness effects, plasmids should be lost: when parasitic (costs outweigh benefits), plasmids should decline due to purifying selection [4, 5 and 6], yet under mutualism (benefits outweigh costs), selection favors the capture of beneficial accessory genes by the chromosome and loss of the costly plasmid backbone [4]. While compensatory evolution can enhance plasmid stability within populations [7, 8, 9, 10, 11, 12, 13, 14 and 15], the propensity for this to occur across the parasitism-mutualism continuum is unknown. We experimentally evolved Pseudomonas fluorescens and its mercury resistance mega-plasmid, pQBR103 [ 16], across an environment-mediated parasitism-mutualism continuum. Compensatory evolution stabilized plasmids by rapidly ameliorating the cost of plasmid carriage in all environments. Genomic analysis revealed that, in both parasitic and mutualistic treatments, evolution repeatedly targeted the gacA/gacS bacterial two-component global regulatory system while leaving the plasmid sequence intact. Deletion of either gacA or gacS was sufficient to completely ameliorate the cost of plasmid carriage. Mutation of gacA/gacS downregulated the expression of ∼17% of chromosomal and plasmid genes and appears to have relieved the translational demand imposed by the plasmid. Chromosomal capture of mercury resistance accompanied by plasmid loss occurred throughout the experiment but very rarely invaded to high frequency, suggesting that rapid compensatory evolution can limit this process. Compensatory evolution can explain the widespread occurrence of plasmids and allows bacteria to retain horizontally acquired plasmids even in environments where their accessory genes are not immediately useful

REFILL: low-cost fillers from quarry waste

Leahill Quarry is located on the coast of SW Ireland (near Adrigole, Co. Cork) in an Upper Devonian gritstone (fine-grained sandstone) sequence that is worked to produce high specification crushed rock aggregates (exported into UK and mainland Europe). The nature of the rock results in large quantities of fines being produced during processing. Leahill Quarry produces about 270,000 tonnes per annum of fines (0-2mm materials) and ‘filler’ (material passing 75m) after processing (washing). In an attempt to find new outlets for Leahill fines, Tarmac Fleming and BGS took part in a three year EC–supported industrial research project called REFILL, which included other UK and Greek partners and was managed overall by MIRO (Mineral Industry Research Organisation). Further details on the project are available by contacting MIRO through their website at: www.miro.co.uk. The REFILL project consisted of various tasks which involved both Tarmac Fleming and BGS. Tasks 1 to 3 involved a study of existing production practices and a programme of technical evaluation of the fines from Leahill and other quarries in Britain. This involved characterization of the mineralogical, physical and chemical properties of the residues. Task 4 was a large-scale continuous test at Leahill Quarry to determine the effectiveness of a novel fluidised bed plant in terms of its ability to extract ‘filler’ from the 0-2mm fines. Tasks 5 to 8 investigated the potential end uses that could incorporate Leahill filler and fines, including an assessment of the available resources of fines, the availability of fines acceptable for particular applications and a review of the market potential for end-uses including asphalt, concrete, artificial soils and non-plastic sub-base materials

Art, Collaboration and Multi-Sensory Approaches in Public Microhistory: Journey with Absent Friends

In this article we reflect upon the many advantages of collaborations between academic historians and artists, as a method for presenting our work, communicating with different audiences and, most importantly, beginning conversations which cause us to think about our research in new and creative ways. We argue that collaboration allows us to rethink the boundaries of expertise, it opens up both historical knowledge and the process of knowledge creation to different audiences in more egalitarian ways, and it provides innovative ways of doing historical research

Rapid compensatory evolution promotes the survival of conjugative plasmids

Conjugative plasmids play a vital role in bacterial adaptation through horizontal gene transfer. Explaining how plasmids persist in host populations however is difficult, given the high costs often associated with plasmid carriage. Compensatory evolution to ameliorate this cost can rescue plasmids from extinction. In a recently published study we showed that compensatory evolution repeatedly targeted the same bacterial regulatory system, GacA/GacS, in populations of plasmid-carrying bacteria evolving across a range of selective environments. Mutations in these genes arose rapidly and completely eliminated the cost of plasmid carriage. Here we extend our analysis using an individual based model to explore the dynamics of compensatory evolution in this system. We show that mutations which ameliorate the cost of plasmid carriage can prevent both the loss of plasmids from the population and the fixation of accessory traits on the bacterial chromosome. We discuss how dependent the outcome of compensatory evolution is on the strength and availability of such mutations and the rate at which beneficial accessory traits integrate on the host chromosome

Bacteriophages limit the existence conditions for conjugative plasmids

Bacteriophages are a major cause of bacterial mortality and impose strong selection on natural bacterial populations, yet their effects on the dynamics of conjugative plasmids have rarely been tested. We combined experimental evolution, mathematical modeling, and individual-based simulations to explain how the ecological and population genetics effects of bacteriophages upon bacteria interact to determine the dynamics of conjugative plasmids and their persistence. The ecological effects of bacteriophages on bacteria are predicted to limit the existence conditions for conjugative plasmids, preventing persistence under weak selection for plasmid accessory traits. Experiments showed that phages drove faster extinction of plasmids in environments where the plasmid conferred no benefit, but they also revealed more complex effects of phages on plasmid dynamics under these conditions, specifically, the temporary maintenance of plasmids at fixation followed by rapid loss. We hypothesized that the population genetic effects of bacteriophages, specifically, selection for phage resistance mutations, may have caused this. Further mathematical modeling and individual-based simulations supported our hypothesis, showing that conjugative plasmids may hitchhike with phage resistance mutations in the bacterial chromosome

The Impact of Mercury Selection and Conjugative Genetic Elements on Community Structure and Resistance Gene Transfer

Carriage of resistance genes can underpin bacterial survival, and by spreading these genes between species, mobile genetic elements (MGEs) can potentially protect diversity within microbial communities. The spread of MGEs could be affected by environmental factors such as selection for resistance, and biological factors such as plasmid host range, with consequences for individual species and for community structure. Here we cultured a focal bacterial strain,Pseudomonas fluorescensSBW25, embedded within a soil microbial community, with and without mercury selection, and with and without mercury resistance plasmids (pQBR57 or pQBR103), to investigate the effects of selection and resistance gene introduction on (1) the focal species; (2) the community as a whole; (3) the spread of the introducedmerresistance operon. We found thatP. fluorescensSBW25 only escaped competitive exclusion by other members of community under mercury selection, even when it did not begin with a mercury resistance plasmid, due to its propensity to acquire resistance from the community by horizontal gene transfer. Mercury pollution had a significant effect on community structure, decreasing alpha diversity within communities while increasing beta diversity between communities, a pattern that was not affected by the introduction of mercury resistance plasmids byP. fluorescensSBW25. Nevertheless, the introducedmerAgene spread to a phylogenetically diverse set of recipients over the 5 weeks of the experiment, as assessed by epicPCR. Our data demonstrates how the effects of MGEs can be experimentally assessed for individual lineages, the wider community, and for the spread of adaptive traits.Peer reviewe

Plasmid carriage can limit bacteria–phage coevolution

Coevolution with bacteriophages is a major selective force shaping bacterial populations and communities. A variety of both environmental and genetic factors has been shown to influence the mode and tempo of bacteria–phage coevolution. Here, we test the effects that carriage of a large conjugative plasmid, pQBR103, had on antagonistic coevolution between the bacterium Pseudomonas fluorescens and its phage, SBW25ϕ2. Plasmid carriage limited bacteria–phage coevolution; bacteria evolved lower phage-resistance and phages evolved lower infectivity in plasmid-carrying compared with plasmid-free populations. These differences were not explained by effects of plasmid carriage on the costs of phage resistance mutations. Surprisingly, in the presence of phages, plasmid carriage resulted in the evolution of high frequencies of mucoid bacterial colonies. Mucoidy can provide weak partial resistance against SBW25ϕ2, which may have limited selection for qualitative resistance mutations in our experiments. Taken together, our results suggest that plasmids can have evolutionary consequences for bacteria that go beyond the direct phenotypic effects of their accessory gene cargo

Conflicting selection alters the trajectory of molecular evolution in a tripartite bacteria-plasmid-phage interaction

Bacteria engage in a complex network of ecological interactions, which includes mobile genetic elements (MGEs) such as phages and plasmids. These elements play a key role in microbial communities as vectors of horizontal gene transfer but can also be important sources of selection for their bacterial hosts. In natural communities bacteria are likely to encounter multiple MGEs simultaneously and conflicting selection among MGEs could alter the bacterial evolutionary response to each MGE. Here we test the effect of interactions with multiple MGEs on bacterial molecular evolution in the tripartite interaction between the bacterium, Pseudomonas fluorescens, the lytic bacteriophage SBW25φ2 and conjugative plasmid, pQBR103, using genome sequencing of experimentally evolved bacteria. We show that, individually, both plasmids and phages impose selection leading to bacterial evolutionary responses that are distinct from bacterial populations evolving without MGEs, but that together, plasmids and phages impose conflicting selection on bacteria, constraining the evolutionary responses observed in pairwise interactions. Our findings highlight the likely difficulties of predicting evolutionary responses to multiple selective pressures from the observed evolutionary responses to each selective pressure alone. Understanding evolution in complex microbial communities comprising many species and MGEs will require that we go beyond studies of pairwise interactions

Viral host-adaptation : insights from evolution experiments with phages

Phages, viral parasites of bacteria, share fundamental features of pathogenic animal and plant viruses and represent a highly tractable empirical model system to understand viral evolution and in particular viral host-adaptation. Phage adaptation to a particular host genotype often results in improved fitness by way of parallel evolution whereby independent lineages hit upon identical adaptive solutions. By contrast, phage adaptation to an evolving host population leads to the evolution of increasing host-range over time and correlated phenotypic and genetic divergence between populations. Phage host-range expansion frequently occurs by a process of stepwise evolution of multiple mutations, and host-shifts are often constrained by mutational availability, pleiotropic costs or ecological conditions

Personalising laboratory medicine in the ‘real world’:assessing clinical utility, by clinical indication, of serum total B12 and Active-B12® (holotranscobalamin) in the diagnosis of vitamin B12 deficiency

BACKGROUND: Assessing the pre- and post-test probability of disease in the context of routine health care is challenging. We wished to study how test performance parameters relating to clinical utility vary by clinical indication in a ‘real-world’ setting. METHODS: The diagnostic accuracy of serum total B(12) and Active-B(12)® (holotranscobalamin) was evaluated in a primary care population, using serum methylmalonic acid as the reference standard. We used electronic requesting to establish the clinical indication for each request. Routine requests from primary care for serum total B(12) were included if creatinine was also measured and estimated glomerular filtration rate was at least 60 mL/min/1.73 m(2). RESULTS: Clinical indications included peripheral neuropathy (n = 168), anaemia (n = 168), cognitive decline (n = 125), suspected dietary deficiency (n = 76), other (n = 362). For peripheral neuropathy, the area under the receiver operator curve ± 95% confidence interval (AUC ± CI) was 0.63 (0.54–0.71) (P = 0.002) for total B(12) and 0.68 (0.60–0.77) (P < 0.0001) for Active-B(12)®. For anaemia, AUC ± CI was 0.56 (0.47–0.66) (P = 0.10) for total B(12) and 0.69 (0.59–0.78) (P < 0.0001) for Active-B(12)®. For cognitive decline, AUC ± CI was 0.54 (0.43–0.65) (P = 0.26) for total B(12) and 0.69 (0.58–0.80) (P = 0.0002) for Active-B(12)®. The pre–post-test change in probability of disease varied by clinical indication. CONCLUSION: Combining diagnostic accuracy studies and electronic testing in a ‘real-world’ setting allows clinical utility to be assessed by clinical indication. Wider application of this would permit more personalised laboratory medicine. In this study, diagnostic performance of total B(12) and Active-B(12)® varied across all indications. Active-B(12)® provided better discrimination, but this may have reflected the cut-offs used