Manipulating multi-level selection in a fungal entomopathogen reveals social conflicts and a method for improving biocontrol traits.

Changes in parasite virulence are commonly expected to lead to trade-offs in other life history traits that can affect fitness. Understanding these trade-offs is particularly important if we want to manipulate the virulence of microbial biological control agents. Theoretically, selection across different spatial scales, i.e. between- and within-hosts, shapes these trade-offs. However, trade-offs are also dependent on parasite biology. Despite their applied importance the evolution of virulence in fungal parasites is poorly understood: virulence can be unstable in culture and commonly fails to increase in simple passage experiments. We hypothesized that manipulating selection intensity at different scales would reveal virulence trade-offs in a fungal pathogen of aphids, Akanthomyces muscarius. Starting with a genetically diverse stock we selected for speed of kill, parasite yield or infectivity by manipulating competition within and between hosts and between-populations of hosts over 7 rounds of infection. We characterized ancestral and evolved lineages by whole genome sequencing and by measuring virulence, growth rate, sporulation and fitness. While several lineages showed increases in virulence, we saw none of the trade-offs commonly found in obligately-killing parasites. Phenotypically similar lineages within treatments often shared multiple single-nucleotide variants, indicating strong convergent evolution. The most dramatic phenotypic changes were in timing of sporulation and spore production in vitro. We found that early sporulation led to reduced competitive fitness but could increase yield of spores on media, a trade-off characteristic of social conflict. Notably, the selection regime with strongest between-population competition and lowest genetic diversity produced the most consistent shift to early sporulation, as predicted by social evolution theory. Multi-level selection therefore revealed social interactions novel to fungi and showed that these biocontrol agents have the genomic flexibility to improve multiple traits-virulence and spore production-that are often in conflict in other parasites.

Antarctica's wilderness fails to capture continent's biodiversity.

Recent assessments of Earth's dwindling wilderness have emphasized that Antarctica is a crucial wilderness in need of protection1,2. Yet human impacts on the continent are widespread3-5, the extent of its wilderness unquantified2 and the importance thereof for biodiversity conservation unknown. Here we assemble a comprehensive record of human activity (approximately 2.7 million records, spanning 200 years) and use it to quantify the extent of Antarctica's wilderness and its representation of biodiversity. We show that 99.6% of the continent's area can still be considered wilderness, but this area captures few biodiversity features. Pristine areas, free from human interference, cover a much smaller area (less than 32% of Antarctica) and are declining as human activity escalates6. Urgent expansion of Antarctica's network of specially protected areas7 can both reverse this trend and secure the continent's biodiversity8-10.

Scaling up ocean conservation through recognition of key biodiversity areas in the Southern Ocean from multispecies tracking data.

Biodiversity is critical for maintaining ecosystem function but is threatened by increasing anthropogenic pressures. In the Southern Ocean, a highly biologically productive region containing many endemic species, proactive management is urgently needed to mitigate increasing pressures from fishing, climate change, and tourism. Site-based conservation is one important tool for managing the negative impacts of human activities on ecosystems. The Key Biodiversity Area (KBA) Standard is a standardized framework used to define sites vital for the persistence of global biodiversity based on criteria and quantitative thresholds. We used tracking data from 14 species of Antarctic and subantarctic seabirds and pinnipeds from the publicly available Retrospective Analysis of Antarctic Tracking Data (RAATD) data set to define KBAs for a diverse suite of marine predators. We used track2kba, an R package that supports identification of KBAs from telemetry data through identification of highly used habitat areas and estimates of local abundance within sites. We compared abundance estimates at each site with thresholds for KBA criteria A1, B1, and D1 (related to globally threatened species, individual geographically restricted species, and demographic aggregations, respectively). We identified 30 potential KBAs for 13 species distributed throughout the Southern Ocean that were vital for each individual species, population, and life-history stage for which they were determined. These areas were identified as highly used by these populations based on observational data and complement the ongoing habitat modeling and bioregionalization work that has been used to prioritize conservation areas in this region. Although further work is needed to identify potential KBAs based on additional current and future data sets, we highlight the benefits of utilizing KBAs as part of a holistic approach to marine conservation, given their significant value as a global conservation tool.

Ampliación de la conservación oceánica por medio del reconocimiento de áreas importantes de biodiversidad en el Océano Antártico a partir de datos de rastreo de varias especies Resumen La biodiversidad es fundamental para mantener la función de los ecosistemas, pero está amenazada por las crecientes presiones antropogénicas. En el Océano Antártico, una región con mucha producción biológica que contiene numerosas especies endémicas, se necesita urgentemente una gestión proactiva para mitigar las crecientes presiones de la pesca, el cambio climático y el turismo. La conservación basada en el sitio es una herramienta importante para gestionar los efectos negativos de las actividades humanas en los ecosistemas. El Estándar de Áreas Clave para la Biodiversidad (ACB) es un marco estandarizado que se utiliza para definir lugares vitales para la persistencia de la biodiversidad mundial con base en criterios y umbrales cuantitativos. Usamos datos del seguimiento de 14 especies de aves marinas y pinnípedos antárticos y sub­antárticos del conjunto de datos públicos Retrospective Analysis of Antarctic Tracking Data (RAATD) para definir las ACB de un conjunto diverso de depredadores marinos. Utilizamos track2kba, un paquete de R que permite la identificación de ACB a partir de datos telemétricos mediante la identificación de áreas de hábitat altamente utilizadas y estimaciones de abundancia local dentro de los sitios. Comparamos las estimaciones de abundancia en cada lugar con los umbrales de los criterios A1, B1 y D1 de las ACB (relacionados con especies amenazadas a nivel mundial, especies individuales restringidas geográficamente y agregaciones demográficas, respectivamente). Identificamos 30 ACB potenciales para 13 especies distribuidas por todo el Océano Antártico que eran vitales para cada especie individual, población y etapa del ciclo biológico para las que se determinaron. Estas áreas fueron identificadas como muy utilizadas por estas poblaciones con base a datos observacionales y complementan el trabajo en curso de modelos del hábitat y biorregionalización que se ha utilizado para priorizar las áreas de conservación en esta región. Aunque es necesario seguir trabajando para identificar posibles ACB basadas en conjuntos de datos adicionales actuales y futuros, destacamos los beneficios de utilizar las ACB como parte de un enfoque holístico de la conservación marina, dado su importante valor como herramienta de conservación global.

The effect of diet composition on the diversity of active gut bacteria and on the growth of Spodoptera exigua (Lepidoptera: Noctuidae).

Gut microbiota plays a functional role in nutrition among several insects. However, the situation is unclear in Lepidoptera. Field studies suggest the microbiome may not be stable and is determined by diet, while in the laboratory, Lepidoptera are routinely reared on diet containing antibiotics with unknown effects on microbial communities. Furthermore, molecular approaches for the characterization of lepidopteran microbiomes rarely describe the metabolically active gut bacteria. The aim of this study was to evaluate how diet and antibiotics affect Spodoptera exigua (Hübner) growth and the diversity and activity of the gut bacteria community. We assessed how alfalfa and wheat germ-based diets affected larval growth, in the presence and absence of streptomycin. Alfalfa diet improved larval growth, pupal mass, and survival, but antibiotic was only beneficial in the wheat germ diet. We observed diet-driven changes in the gut bacterial communities. In the active community, the alfalfa colony was dominated by Enterococcus and Rhodococcus whereas in the wheat germ colony, only Enterococcus was present. In contrast, spore-forming Bacilli species were very common members of the DNA community. In both cases, streptomycin had a selective effect on the relative abundance of the taxa present. Our study highlights the importance of characterizing both the diversity and activity of the gut microbiota community. DNA-derived communities may include environmental DNA, spores, or non-viable bacteria, while RNA-derived communities are more likely to give an accurate representation of the diversity of active members that are potentially directly involved in the metabolic processes of the host.

Decolonizing drug-resistant E. coli with phage and probiotics: breaking the frequency-dependent dominance of residents.

Widespread antibiotic resistance in commensal bacteria creates a persistent challenge for human health. Resident drug-resistant microbes can prevent clinical interventions, colonize wounds post-surgery, pass resistance traits to pathogens or move to more harmful niches following routine interventions such as catheterization. Accelerating the removal of resistant bacteria or actively decolonizing particular lineages from hosts could therefore have a number of long-term benefits. However, removing resident bacteria via competition with probiotics, for example, poses a number of ecological challenges. Resident microbes are likely to have physiological and numerical advantages and competition based on bacteriocins or other secreted antagonists is expected to give advantages to the dominant partner, via positive frequency dependence. Since a narrow range of Escherichia coli genotypes (primarily those belonging to the clonal group ST131) cause a significant proportion of multidrug-resistant infections, this group presents a promising target for decolonization with bacteriophage, as narrow-host-range viral predation could lead to selective removal of particular genotypes. In this study we tested how a combination of an ST131-specific phage and competition from the well-known probiotic E. coli Nissle strain could displace E. coli ST131 under aerobic and anaerobic growth conditions in vitro. We showed that the addition of phage was able to break the frequency-dependent advantage of a numerically dominant ST131 isolate. Moreover, the addition of competing E. coli Nissle could improve the ability of phage to suppress ST131 by two orders of magnitude. Low-cost phage resistance evolved readily in these experiments and was not inhibited by the presence of a probiotic competitor. Nevertheless, combinations of phage and probiotic produced stable long-term suppression of ST131 over multiple transfers and under both aerobic and anaerobic growth conditions. Combinations of phage and probiotic therefore have real potential for accelerating the removal of drug-resistant commensal targets.

Group Selection as a Basis for Screening Mutagenized Libraries of Public Goods (Bacillus thuringiensis Cry Toxins).

The pesticidal toxins of Bacillus thuringiensis (Bt) supply the active proteins for genetically modified insect-resistant crops. There is therefore keen interest in finding new toxins, or improving known toxins, in order to increase the mortality of various targets. The production and screening of large libraries of mutagenized toxins are among the means of identifying improved toxins. Since Cry toxins are public goods, and do not confer advantages to producers in competition, conventional directed evolution approaches cannot be used here. Instead, thousands of individual mutants have to be sequenced and assayed individually, a costly and time-consuming process. In this study, we tested a group selection-based approach that could be used to screen an uncharacterized pool of Cry toxin mutants. This involved selecting for infectivity between subpopulations of Bt clones within metapopulations of infected insects in three rounds of passage. We also tested whether additional mutagenesis from exposure to ethyl methanesulfonate could increase infectivity or supply additional Cry toxin diversity during passage. Sequencing of pools of mutants at the end of selection showed that we could effectively screen out Cry toxin variants that had reduced toxicity with our group selection approach. The addition of extra mutagenesis during passage decreased the efficiency of selection for infectivity and did not produce any additional novel toxin diversity. Toxins with loss-of-function mutations tend to dominate mutagenized libraries, and so a process for screening out these mutants without time-consuming sequencing and characterization steps could be beneficial when applied to larger libraries. IMPORTANCE Insecticidal toxins from the bacterium Bacillus thuringiensis are widely exploited in genetically modified plants. This application creates a demand for novel insecticidal toxins that can be used to better manage resistant pests or control new or recalcitrant target species. An important means of producing novel toxins is via high-throughput mutagenesis and screening of existing toxins, a lengthy and resource-intensive process. This study describes the development and testing of an efficient means of screening a test library of mutagenized insecticidal toxins. Here, we showed that it is possible to screen out loss-of-function mutations with low infectivity within a pool without the need to characterize and sequence each mutant individually. This has the potential to improve the efficiency of processes used to identify novel proteins.

Climate change drives expansion of Antarctic ice-free habitat.

Antarctic terrestrial biodiversity occurs almost exclusively in ice-free areas that cover less than 1% of the continent. Climate change will alter the extent and configuration of ice-free areas, yet the distribution and severity of these effects remain unclear. Here we quantify the impact of twenty-first century climate change on ice-free areas under two Intergovernmental Panel on Climate Change (IPCC) climate forcing scenarios using temperature-index melt modelling. Under the strongest forcing scenario, ice-free areas could expand by over 17,000 km2 by the end of the century, close to a 25% increase. Most of this expansion will occur in the Antarctic Peninsula, where a threefold increase in ice-free area could drastically change the availability and connectivity of biodiversity habitat. Isolated ice-free areas will coalesce, and while the effects on biodiversity are uncertain, we hypothesize that they could eventually lead to increasing regional-scale biotic homogenization, the extinction of less-competitive species and the spread of invasive species.

Signatures of selection in core and accessory genomes indicate different ecological drivers of diversification among Bacillus cereus clades.

Bacterial clades are often ecologically distinct, despite extensive horizontal gene transfer (HGT). How selection works on different parts of bacterial pan-genomes to drive and maintain the emergence of clades is unclear. Focusing on the three largest clades in the diverse and well-studied Bacillus cereus sensu lato group, we identified clade-specific core genes (present in all clade members) and then used clade-specific allelic diversity to identify genes under purifying and diversifying selection. Clade-specific accessory genes (present in a subset of strains within a clade) were characterized as being under selection using presence/absence in specific clades. Gene ontology analyses of genes under selection revealed that different gene functions were enriched in different clades. Furthermore, some gene functions were enriched only amongst clade-specific core or accessory genomes. Genes under purifying selection were often clade-specific, while genes under diversifying selection showed signs of frequent HGT. These patterns are consistent with different selection pressures acting on both the core and the accessory genomes of different clades and can lead to ecological divergence in both cases. Examining variation in allelic diversity allows us to uncover genes under clade-specific selection, allowing ready identification of strains and their ecological niche.

E-cigarette addiction and harm perception: Does initiation flavor choice matter?

INTRODUCTION: The 21st century was marked by a dramatic increase in adolescent e-cigarette use in the United States (US). The popularity of non-traditional flavor types, including fruit and pastry, is thought to contribute toward growing product use nationally, leading to a variety of federal and state regulations limiting the use of non-traditional flavors in the US. The relationship between flavor type and increased adolescent use suggests a possible link between flavor use and addiction and harm perception. This study assessed if the flavor type used when initiating e-cigarette use predicted addiction and harm perceptions. METHODS: The study utilized data from the multi-wave youth Population Assessment of Tobacco Health Study. It explored the impact initiating e-cigarette use with traditional versus non-traditional flavor types among cigarette users on the outcome variables: e-cigarette addiction and harm perception. Both e-cigarette addiction and harm perception were measured using self-report, Likert scale questionnaires. Descriptive statistics characterized the study variables and linear regression analyses performed to test whether flavor initiation type is associated with addiction and harm perception. RESULTS: The study sample consisted of 1,043 youth (weighted N = 1,873,617) aged 12 to 17 years who reported at least one instance of e-cigarette use. After adjusting for age, age of onset, sex, race and annual household income there was no statistically significant difference in addiction levels between those initiating with traditional versus non-traditional flavors (p = 0.294). Similarly, traditional versus non-traditional flavor initiation did not show a statistically significant difference in adolescent e-cigarette harm perceptions (p = 0.601). CONCLUSIONS: Traditionally flavored e-cigarette initiation produces similar risk for addiction and harm perceptions as non-traditionally flavored initiation. These findings suggest that banning non-traditional flavors alone may be ineffective in curbing e-cigarette addiction and harm perception. Additional research is needed to better understand which e-cigarette product characteristics and behaviors may be associated with greater addiction and reduced harm perceptions.

Passage and the evolution of virulence in invertebrate pathogens: Fundamental and applied perspectives.

Understanding the ecological and genetic factors that determine the evolution of virulence has broad value for invertebrate pathology. In addition to helping us understand the fundamental biology of our study organisms this body of theory has important applications in microbial biocontrol. Experimental tests of virulence theory are often carried out in invertebrate models and yet theory rarely informs applied passage experiments that aim to increase or maintain virulence. This review summarizes recent progress in this field with a focus on work most relevant to biological control: the virulence of invertebrate pathogens that are 'obligate killers' and which require cadavers for the production of infectious propagules. We discuss recent theory and fundamental and applied experimental evolution with bacteria, fungi, baculoviruses and nematodes. While passage experiments using baculoviruses have a long history of producing isolates with increased virulence, studies with other pathogens have not been so successful. Recent passage experiments that have applied evolution of virulence frameworks based on cooperation (kin selection) have produced novel methods and promising mutants with increased killing power. Evolution of virulence theory can provide plausible explanations for the varied results of passage experiments as well as a predictive framework for improving artificial selection.

Function is a better predictor of plant rhizosphere community membership than 16S phylogeny.

Rhizobacterial communities are important for plant health but we still have limited understanding of how they are constructed or how they can be manipulated. High-throughput 16S rRNA sequencing provides good information on taxonomic composition but remains an unreliable proxy for phenotypes. In this study, we tested the hypothesis that experimentally observed functional traits would be better predictors of community membership than phylogenetic origin. To test this hypothesis, we sampled communities on four plant species grown in two soil types and characterized 593 bacterial isolates in terms of antibiotic susceptibility, carbon metabolism, resource use and plant growth-promoting traits. In support of our hypothesis we found that three of the four plant species had phylogenetically diverse, but functionally constrained communities. Notably, communities did not grow best on complex media mimicking their host of origin but were distinguished by variation in overall growth characteristics (copiotrophy/oligotrophy) and antibiotic susceptibility. These data, combined with variation in phylogenetic structure, suggest that different classes of traits (antagonistic competition or resource-based) are more important in different communities. This culture-based approach supports and complements the findings of a previous high-throughput 16S rRNA analysis of this experiment and provides functional insights into the patterns observed with culture-independent methods.

Combating ecosystem collapse from the tropics to the Antarctic.

Globally, collapse of ecosystems-potentially irreversible change to ecosystem structure, composition and function-imperils biodiversity, human health and well-being. We examine the current state and recent trajectories of 19 ecosystems, spanning 58° of latitude across 7.7 M km2 , from Australia's coral reefs to terrestrial Antarctica. Pressures from global climate change and regional human impacts, occurring as chronic 'presses' and/or acute 'pulses', drive ecosystem collapse. Ecosystem responses to 5-17 pressures were categorised as four collapse profiles-abrupt, smooth, stepped and fluctuating. The manifestation of widespread ecosystem collapse is a stark warning of the necessity to take action. We present a three-step assessment and management framework (3As Pathway Awareness, Anticipation and Action) to aid strategic and effective mitigation to alleviate further degradation to help secure our future.

Ugandan stakeholder hopes and concerns about gene drive mosquitoes for malaria control: new directions for gene drive risk governance.

BACKGROUND: The African Union's High-Level Panel on Emerging Technologies identified gene drive mosquitoes as a priority technology for malaria elimination. The first field trials are expected in 5-10 years in Uganda, Mali or Burkina Faso. In preparation, regional and international actors are developing risk governance guidelines which will delineate the framework for identifying and evaluating risks. Scientists and bioethicists have called for African stakeholder involvement in these developments, arguing the knowledge and perspectives of those people living in malaria-afflicted countries is currently missing. However, few African stakeholders have been involved to date, leaving a knowledge gap about the local social-cultural as well as ecological context in which gene drive mosquitoes will be tested and deployed. This study investigates and analyses Ugandan stakeholders' hopes and concerns about gene drive mosquitoes for malaria control and explores the new directions needed for risk governance. METHODS: This qualitative study draws on 19 in-depth semi-structured interviews with Ugandan stakeholders in 2019. It explores their hopes for the technology and the risks they believed pertinent. Coding began at a workshop and continued through thematic analysis. RESULTS: Participants' hopes and concerns for gene drive mosquitoes to address malaria fell into three themes: (1) ability of gene drive mosquitoes to prevent malaria infection; (2) impacts of gene drive testing and deployment; and, (3) governance. Stakeholder hopes fell almost exclusively into the first theme while concerns were spread across all three. The study demonstrates that local stakeholders are able and willing to contribute relevant and important knowledge to the development of risk frameworks. CONCLUSIONS: International processes can provide high-level guidelines, but risk decision-making must be grounded in the local context if it is to be robust, meaningful and legitimate. Decisions about whether or not to release gene drive mosquitoes as part of a malaria control programme will need to consider the assessment of both the risks and the benefits of gene drive mosquitoes within a particular social, political, ecological, and technological context. Just as with risks, benefits-and importantly, the conditions that are necessary to realize them-must be identified and debated in Uganda and its neighbouring countries.

Managing ecosystems in a sea of uncertainty: invasive species management and assisted colonizations.

Managing ecosystems in the face of complex species interactions, and the associated uncertainty, presents a considerable ecological challenge. Altering those interactions via actions such as invasive species management or conservation translocations can result in unintended consequences, supporting the need to be able to make more informed decisions in the face of this uncertainty. We demonstrate the utility of ecosystem models to reduce uncertainty and inform future ecosystem management. We use Phillip Island, Australia, as a case study to investigate the impacts of two invasive species management options and consider whether a critically endangered mammal is likely to establish a population in the presence of invasive species. Qualitative models are used to determine the effects of apex predator removal (feral cats) and invasive prey removal (rabbits, rats, and mice). We extend this approach using Ensemble Ecosystem Models to consider how suppression, rather than eradication influences the species community; and consider whether an introduction of the critically endangered eastern barred bandicoot is likely to be successful in the presence of invasive species. Our analysis revealed the potential for unintended outcomes associated with feral cat control operations, with rats and rabbits expected to increase in abundance. A strategy based on managing prey species appeared to have the most ecosystem-wide benefits, with rodent control showing more favorable responses than a rabbit control strategy. Eastern barred bandicoots were predicted to persist under all feral cat control levels (including no control). Managing ecosystems is a complex and imprecise process. However, qualitative modeling and ensemble ecosystem modeling address uncertainty and are capable of improving and optimizing management practices. Our analysis shows that the best conservation outcomes may not always be associated with the top-down control of apex predators, and land managers should think more broadly in relation to managing bottom-up processes as well. Challenges faced in continuing to conserve biodiversity mean new, bolder, conservation actions are needed. We suggest that endangered species are capable of surviving in the presence of feral cats, potentially opening the door for more conservation translocations.

Strong Environment-Genotype Interactions Determine the Fitness Costs of Antibiotic Resistance In Vitro and in an Insect Model of Infection.

The acquisition of antibiotic resistance commonly imposes fitness costs, a reduction in the fitness of bacteria in the absence of drugs. These costs have been quantified primarily using in vitro experiments and a small number of in vivo studies in mice, and it is commonly assumed that these diverse methods are consistent. Here, we used an insect model of infection to compare the fitness costs of antibiotic resistance in vivo to those in vitro Experiments explored diverse mechanisms of resistance in a Gram-positive pathogen, Bacillus thuringiensis, and a Gram-negative intestinal symbiont, Enterobacter cloacae Rifampin resistance in B. thuringiensis showed fitness costs that were typically elevated in vivo, although these were modulated by genotype-environment interactions. In contrast, resistance to cefotaxime via derepression of AmpC ß-lactamase in E. cloacae resulted in no detectable costs in vivo or in vitro, while spontaneous resistance to nalidixic acid, and carriage of the IncP plasmid RP4, imposed costs that increased in vivo Overall, fitness costs in vitro were a poor predictor of fitness costs in vivo because of strong genotype-environment interactions throughout this study. Insect infections provide a cheap and accessible means of assessing the fitness consequences of resistance mutations, data that are important for understanding the evolution and spread of resistance. This study emphasizes that the fitness costs imposed by particular mutations or different modes of resistance are extremely variable and that only a subset of these mutations is likely to be prevalent outside the laboratory.

Shifts along the parasite-mutualist continuum are opposed by fundamental trade-offs.

Theory suggests that symbionts can readily evolve more parasitic or mutualistic strategies with respect to hosts. However, many symbionts have stable interactions with hosts that improve nutrient assimilation or confer protection from pathogens. We explored the potential for evolution of increased parasitism or decreased parasitism and mutualism in a natural gut symbiosis between larvae of Plutella xylostella and the microbe Enterobacter cloacae. We focused on interactions with the pathogen, Bacillus thuringiensis: selecting for parasitism in terms of facilitating pathogen infection, or increased mutualism in terms of host protection. Selection for parasitism led to symbionts increasing pathogen-induced mortality but reduced their competitive ability with pathogens and their in vitro growth rates. Symbionts did not evolve to confer protection from pathogens. However, several lineages evolved reduced parasitism, primarily in terms of moderating impacts on host growth, potentially because prudence pays dividends through increased host size. Overall, the evolution of increased parasitism was achievable but was opposed by trade-offs likely to reduce fitness. The evolution of protection may not have occurred because suppressing growth of B. thuringiensis in the gut might provide only weak protection or because evolution towards protective interactions was opposed by the loss of competitive fitness in symbionts.

Bacteria from natural populations transfer plasmids mostly towards their kin.

Plasmids play a key role in microbial ecology and evolution, yet the determinants of plasmid transfer rates are poorly understood. Particularly, interactions between donor hosts and potential recipients are understudied. Here, we investigate the importance of genetic similarity between naturally co-occurring Escherichia coli isolates in plasmid transfer. We uncover extensive variability, spanning over five orders of magnitude, in the ability of isolates to donate and receive two different plasmids, R1 and RP4. Overall, transfer is strongly biased towards clone-mates, but not correlated to genetic distance when donors and recipients are not clone-mates. Transfer is limited by the presence of a functional restriction-modification system in recipients, suggesting sharing of strain-specific defence systems contributes to bias towards kin. Such restriction of transfer to kin sets the stage for longer-term coevolutionary interactions leading to mutualism between plasmids and bacterial hosts in natural communities.

Crystal toxins and the volunteer's dilemma in bacteria.

The growth and virulence of the bacteria Bacillus thuringiensis depend on the production of Cry toxins, which are used to perforate the gut of its host. Successful invasion of the host relies on producing a threshold amount of toxin, after which there is no benefit from producing more toxin. Consequently, the production of Cry toxin appears to be a different type of social problem compared with the public goods scenarios that bacteria usually encounter. We show that selection for toxin production is a volunteer's dilemma. We make specific predictions that (a) selection for toxin production depends upon an interplay between the number of bacterial cells that each host ingests and the genetic relatedness between those cells; (b) cheats that do not produce toxin gain an advantage when at low frequencies, and at high bacterial density, allowing them to be maintained in a population alongside toxin-producing cells. More generally, our results emphasize the diversity of the social games that bacteria play.

Targeting antibiotic resistant bacteria with phage reduces bacterial density in an insect host.

Phage therapy is attracting growing interest among clinicians as antibiotic resistance continues becoming harder to control. However, clinical trials and animal model studies on bacteriophage treatment are still scarce and results on the efficacy vary. Recent research suggests that using traditional antimicrobials in concert with phage could have desirable synergistic effects that hinder the evolution of resistance. Here, we present a novel insect gut model to study phage-antibiotic interaction in a system where antibiotic resistance initially exists in very low frequency and phage specifically targets the resistance bearing cells. We demonstrate that while phage therapy could not reduce the frequency of target bacteria in the population during positive selection by antibiotics, it alleviated the antibiotic induced blooming by lowering the overall load of resistant cells. The highly structured gut environment had pharmacokinetic effects on both phage and antibiotic dynamics compared with in vitro: antibiotics did not reduce the overall amount of bacteria, demonstrating a simple turnover of gut microbiota from non-resistant to resistant population with little cost. The results imply moderate potential for using phage as an aid to target antibiotic resistant gut infections, and question the usefulness of in vitro inferences.

Lineage-specific plasmid acquisition and the evolution of specialized pathogens in Bacillus thuringiensis and the Bacillus cereus group.

Bacterial plasmids can vary from small selfish genetic elements to large autonomous replicons that constitute a significant proportion of total cellular DNA. By conferring novel function to the cell, plasmids may facilitate evolution but their mobility may be opposed by co-evolutionary relationships with chromosomes or encouraged via the infectious sharing of genes encoding public goods. Here, we explore these hypotheses through large-scale examination of the association between plasmids and chromosomal DNA in the phenotypically diverse Bacillus cereus group. This complex group is rich in plasmids, many of which encode essential virulence factors (Cry toxins) that are known public goods. We characterized population genomic structure, gene content and plasmid distribution to investigate the role of mobile elements in diversification. We analysed coding sequence within the core and accessory genome of 190 B. cereus group isolates, including 23 novel sequences and genes from 410 reference plasmid genomes. While cry genes were widely distributed, those with invertebrate toxicity were predominantly associated with one sequence cluster (clade 2) and phenotypically defined Bacillus thuringiensis. Cry toxin plasmids in clade 2 showed evidence of recent horizontal transfer and variable gene content, a pattern of plasmid segregation consistent with transfer during infectious cooperation. Nevertheless, comparison between clades suggests that co-evolutionary interactions may drive association between plasmids and chromosomes and limit wider transfer of key virulence traits. Proliferation of successful plasmid and chromosome combinations is a feature of specialized pathogens with characteristic niches (Bacillus anthracis, B. thuringiensis) and has occurred multiple times in the B. cereus group.