Browsing by Author "Sousa, Ana"
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- Evolution of commensal bacteria in the intestinal tract of micePublication . Sousa, Ana; Frazão, Nelson; Ramiro, Ricardo S; Gordo, IsabelHundreds of different bacterial species inhabit our intestines and contribute to our health status, with significant loss of species diversity typically observed in disease conditions. Within each microbial species a great deal of diversity is hidden and such intra-specific variation is also key to the proper homeostasis between the host and its microbial inhabitants. Indeed, it is at this level that new mechanisms of antibiotic resistance emerge and pathogenic characteristics evolve. Yet, our knowledge on intra-species variation in the gut is still limited and an understanding of the evolutionary mechanisms acting on it is extremely reduced. Here we review recent work that has begun to reveal that adaptation of commensal bacteria to the mammalian intestine may be fast and highly repeatable, and that the time scales of evolutionary and ecological change can be very similar in these ecosystems.
- Multiple Resistance at No Cost: Rifampicin and Streptomycin a Dangerous Liaison in the Spread of Antibiotic ResistancePublication . Durão, Paulo; Trindade, Sandra; Sousa, Ana; Gordo, IsabelEvidence is mounting that epistasis is widespread among mutations. The cost of carrying two deleterious mutations, or the advantage of acquiring two beneficial alleles, is typically lower that the sum of their individual effects. Much less is known on epistasis between beneficial and deleterious mutations, even though this is key to the amount of genetic hitchhiking that may occur during evolution. This is particularly important in the context of antibiotic resistance: Most resistances are deleterious, but some can be beneficial and remarkably rifampicin resistance can emerge de novo in populations evolving without antibiotics. Here we show pervasive positive pairwise epistasis on Escherichia coli fitness between beneficial mutations, which confer resistance to rifampicin, and deleterious mutations, which confer resistance to streptomycin. We find that 65% of double resistant strains outcompete sensitive bacteria in an environment devoid of antibiotics. Weak beneficial mutations may therefore overcome strong deleterious mutations and can even render double mutants strong competitors.
- A Mutational Hotspot and Strong Selection Contribute to the Order of Mutations Selected for during Escherichia coli Adaptation to the GutPublication . Lourenço, Marta; Ramiro, Ricardo S; Güleresi, Daniela; Barroso-Batista, João; Xavier, Karina B; Gordo, Isabel; Sousa, AnaThe relative role of drift versus selection underlying the evolution of bacterial species within the gut microbiota remains poorly understood. The large sizes of bacterial populations in this environment suggest that even adaptive mutations with weak effects, thought to be the most frequently occurring, could substantially contribute to a rapid pace of evolutionary change in the gut. We followed the emergence of intra-species diversity in a commensal Escherichia coli strain that previously acquired an adaptive mutation with strong effect during one week of colonization of the mouse gut. Following this first step, which consisted of inactivating a metabolic operon, one third of the subsequent adaptive mutations were found to have a selective effect as high as the first. Nevertheless, the order of the adaptive steps was strongly affected by a mutational hotspot with an exceptionally high mutation rate of 10-5. The pattern of polymorphism emerging in the populations evolving within different hosts was characterized by periodic selection, which reduced diversity, but also frequency-dependent selection, actively maintaining genetic diversity. Furthermore, the continuous emergence of similar phenotypes due to distinct mutations, known as clonal interference, was pervasive. Evolutionary change within the gut is therefore highly repeatable within and across hosts, with adaptive mutations of selection coefficients as strong as 12% accumulating without strong constraints on genetic background. In vivo competitive assays showed that one of the second steps (focA) exhibited positive epistasis with the first, while another (dcuB) exhibited negative epistasis. The data shows that strong effect adaptive mutations continuously recur in gut commensal bacterial species.
- Potential for adaptation overrides cost of resistancePublication . Moura de Sousa, Jorge; Sousa, Ana; Bourgard, Catarina; Gordo, IsabelTo investigate the cost of antibiotic resistance versus the potential for resistant clones to adapt in maintaining polymorphism for resistance. Materials & methods: Experimental evolution of Escherichia coli carrying different resistance alleles was performed under an environment devoid of antibiotics and evolutionary parameters estimated from their frequencies along time. Results & conclusion: Costly resistance mutations were found to coexist with lower cost resistances for hundreds of generations, contrary to the hypothesis that the cost of a resistance dictates its extinction. Estimated evolutionary parameters for the different resistance backgrounds suggest a higher adaptive potential of clones with costly antibiotic resistance mutations, overriding their initial cost of resistance and allowing their maintenance in the absence of drugs.
- Recurrent Reverse Evolution Maintains Polymorphism after Strong Bottlenecks in Commensal Gut BacteriaPublication . Sousa, Ana; Ramiro, Ricardo S.; Barroso-Batista, João; Güleresi, Daniela; Lourenço, Marta; Gordo, IsabelThe evolution of new strains within the gut ecosystem is poorly understood. We used a natural but controlled system to follow the emergence of intraspecies diversity of commensal Escherichia coli, during three rounds of adaptation to the mouse gut (∼1,300 generations). We previously showed that, in the first round, a strongly beneficial phenotype (loss-of-function for galactitol consumption; gat-negative) spread to >90% frequency in all colonized mice. Here, we show that this loss-of-function is repeatedly reversed when a gat-negative clone colonizes new mice. The regain of function occurs via compensatory mutation and reversion, the latter leaving no trace of past adaptation. We further show that loss-of-function adaptive mutants reevolve, after colonization with an evolved gat-positive clone. Thus, even under strong bottlenecks a regime of strong-mutation-strong-selection dominates adaptation. Coupling experiments and modeling, we establish that reverse evolution recurrently generates two coexisting phenotypes within the microbiota that can or not consume galactitol (gat-positive and gat-negative, respectively). Although the abundance of the dominant strain, the gat-negative, depends on the microbiota composition, gat-positive abundance is independent of the microbiota composition and can be precisely manipulated by supplementing the diet with galactitol. These results show that a specific diet is able to change the abundance of specific strains. Importantly, we find polymorphism for these phenotypes in indigenous Enterobacteria of mice and man. Our results demonstrate that natural selection can greatly overwhelm genetic drift at structuring the strain diversity of gut commensals and that competition for limiting resources may be a key mechanism for maintaining polymorphism in the gut.
- The Genetic Basis of Escherichia coli Pathoadaptation to MacrophagesPublication . Miskinyte, Migla; Sousa, Ana; Ramiro, Ricardo S.; de Sousa, Jorge A. Moura; Kotlinowski, Jerzy; Caramalho, Iris; Magalhães, Sara; Soares, Miguel P.; Gordo, IsabelAntagonistic interactions are likely important driving forces of the evolutionary process underlying bacterial genome complexity and diversity. We hypothesized that the ability of evolved bacteria to escape specific components of host innate immunity, such as phagocytosis and killing by macrophages (MΦ), is a critical trait relevant in the acquisition of bacterial virulence. Here, we used a combination of experimental evolution, phenotypic characterization, genome sequencing and mathematical modeling to address how fast, and through how many adaptive steps, a commensal Escherichia coli (E. coli) acquire this virulence trait. We show that when maintained in vitro under the selective pressure of host MΦ commensal E. coli can evolve, in less than 500 generations, virulent clones that escape phagocytosis and MΦ killing in vitro, while increasing their pathogenicity in vivo, as assessed in mice. This pathoadaptive process is driven by a mechanism involving the insertion of a single transposable element into the promoter region of the E. coli yrfF gene. Moreover, transposition of the IS186 element into the promoter of Lon gene, encoding an ATP-dependent serine protease, is likely to accelerate this pathoadaptive process. Competition between clones carrying distinct beneficial mutations dominates the dynamics of the pathoadaptive process, as suggested from a mathematical model, which reproduces the observed experimental dynamics of E. coli evolution towards virulence. In conclusion, we reveal a molecular mechanism explaining how a specific component of host innate immunity can modulate microbial evolution towards pathogenicity.