Adaptation of the Mycobacterium tuberculosis transcriptome to biofilm growth.

Mycobacterium tuberculosis (M. tb), the causative agent of tuberculosis (TB), is a leading global cause of death from infectious disease. Biofilms are increasingly recognized as a relevant growth form during M. tb infection and may impede treatment by enabling bacterial drug and immune tolerance. M. tb has a complicated regulatory network that has been well-characterized for many relevant disease states, including dormancy and hypoxia. However, despite its importance, our knowledge of the genes and pathways involved in biofilm formation is limited. Here we characterize the biofilm transcriptomes of fully virulent clinical isolates and find that the regulatory systems underlying biofilm growth vary widely between strains and are also distinct from regulatory programs associated with other environmental cues. We used experimental evolution to investigate changes to the transcriptome during adaptation to biofilm growth and found that the application of a uniform selection pressure resulted in loss of strain-to-strain variation in gene expression, resulting in a more uniform biofilm transcriptome. The adaptive trajectories of transcriptomes were shaped by the genetic background of the M. tb population leading to convergence on a sub-lineage specific transcriptome. We identified widespread upregulation of non-coding RNA (ncRNA) as a common feature of the biofilm transcriptome and hypothesize that ncRNA function in genome-wide modulation of gene expression, thereby facilitating rapid regulatory responses to new environments. These results reveal a new facet of the M. tb regulatory system and provide valuable insight into how M. tb adapts to new environments.

Adaptation of the Mycobacterium tuberculosis transcriptome to biofilm growth.

Mycobacterium tuberculosis ( M. tb ), the causative agent of tuberculosis (TB), is a leading global cause of death from infectious disease. Biofilms are increasingly recognized as a relevant growth form during M. tb infection and may impede treatment by enabling bacterial drug and immune tolerance. M. tb has a complicated regulatory network that has been well-characterized for many relevant disease states, including dormancy and hypoxia. However, despite its importance, our knowledge of the genes and pathways involved in biofilm formation is limited. Here we characterize the biofilm transcriptomes of fully virulent clinical isolates and find that the regulatory systems underlying biofilm growth vary widely between strains and are also distinct from regulatory programs associated with other environmental cues. We used experimental evolution to investigate changes to the transcriptome during adaptation to biofilm growth and found that the application of a uniform selection pressure resulted in loss of strain-to-strain variation in gene expression, resulting in a more uniform biofilm transcriptome. The adaptive trajectories of transcriptomes were shaped by the genetic background of the M. tb population leading to convergence on a sub-lineage specific transcriptome. We identified widespread upregulation of non-coding RNA (ncRNA) as a common feature of the biofilm transcriptome and hypothesize that ncRNA function in genome-wide modulation of gene expression, thereby facilitating rapid regulatory responses to new environments. These results reveal a new facet of the M. tb regulatory system and provide valuable insight into how M. tb adapts to new environments. Importance: Understanding mechanisms of resistance and tolerance in Mycobacterium tuberculosis ( M. tb ) can help us develop new treatments that capitalize on M. tb 's vulnerabilities. Here we used transcriptomics to study both the regulation of biofilm formation in clinical isolates as well as how those regulatory systems adapt to new environments. We find that closely related clinical populations have diverse strategies for growth under biofilm conditions, and that genetic background plays a large role in determining the trajectory of evolution. These results have implications for future treatment strategies that may be informed by our knowledge of the evolutionary constraints on strain(s) from an individual infection. This work provides new information about the mechanisms of biofilm formation in M. tb and outlines a framework for population level approaches for studying bacterial adaptation.

Rapid adaptation of a complex trait during experimental evolution of Mycobacterium tuberculosis.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (M. tb), is a leading cause of death due to infectious disease. TB is not traditionally associated with biofilms, but M. tb biofilms are linked with drug and immune tolerance and there is increasing recognition of their contribution to the recalcitrance of TB infections. Here, we used M. tb experimental evolution to investigate this complex phenotype and identify candidate loci controlling biofilm formation. We identified novel candidate loci, adding to our understanding of the genetic architecture underlying M. tb biofilm development. Under selective pressure to grow as a biofilm, regulatory mutations rapidly swept to fixation and were associated with changes in multiple traits, including extracellular matrix production, cell size, and growth rate. Genetic and phenotypic paths to enhanced biofilm growth varied according to the genetic background of the parent strain, suggesting that epistatic interactions are important in M. tb adaptation to changing environments.

In many environments, bacteria live together in structures called biofilms. Cells in biofilms coordinate with each other to protect the group and allow it to survive difficult conditions. Mycobacterium tuberculosis, the bacterium that causes tuberculosis, forms biofilms when it infects the human body. Biofilms make the infection a lot more difficult to treat, which may be one of the reasons why tuberculosis is the deadliest bacterial infection in the world. Bacteria evolve rapidly over the course of a single infection, but bacteria forming biofilms evolve differently to bacteria living alone. This evolution happens through mutations to the bacterial DNA, which can be small (a single base in a DNA sequence changes to a different base) or larger changes (such as the deletion or insertion of several bases). Smith, Youngblom et al. studied the evolution of tuberculosis growing in biofilms in the lab. As the bacteria evolved, they tended to form thicker biofilms, an effect linked to 14 mutations involving single base DNA changes and four larger ones. Most of the changes were in regulatory regions of DNA, which control whether genes are 'read' by cells to produce proteins. These regions often change more though evolution than regions coding for proteins, because they have a coordinated effect on a group of related genes rather than randomly altering individual genes. Smith, Youngblom et al. also showed that biofilms made from different strains of tuberculosis evolved in different ways. Smith Youngblom et al.'s findings provide more information regarding how bacteria adapt to living in biofilms, which may reveal new ways to control them. This could have applications in water treatment, food production and healthcare. Learning how to treat bacteria growing in biofilms could also improve the outcomes for patients infected with tuberculosis.

Epidemiological and genomic determinants of tuberculosis outbreaks in First Nations communities in Canada.

BACKGROUND: In Canada, tuberculosis disproportionately affects foreign-born and First Nations populations. Within First Nations' peoples, a high proportion of cases occur in association with outbreaks. Tuberculosis transmission in the context of outbreaks is thought to result from the convergence of several factors including characteristics of the cases, contacts, the environment, and the pathogen. METHODS: We examined the epidemiological and genomic determinants of two well-characterized tuberculosis outbreaks attributed to two super-spreaders among First Nations in the province of Alberta. These outbreaks were associated with two distinct DNA fingerprints (restriction fragment-length polymorphisms or RFLPs 0.0142 and 0.0728). We compared outbreak isolates with endemic isolates not spatio-temporarily linked to outbreak cases. We extracted epidemiological variables pertaining to tuberculosis cases and contacts from individual public health records and the provincial tuberculosis registry. We conducted group analyses using parametric and non-parametric statistical tests. We carried out whole-genome sequencing and bioinformatic analysis using validated protocols. RESULTS: We observed differences between outbreak and endemic groups in the mean number of total and child-aged contacts and the number of contacts with new positive and converted tuberculin skin tests in all group comparisons (p < 0.05). Differences were also detected in the proportion of cases with cavitation on a chest radiograph and the mean number of close contacts in selected group comparisons (p < 0.02). A phylogenetic network analysis of whole-genome sequencing data indicated that most outbreak and endemic strains were closely related to the source case for the 0.0142 fingerprint. For the 0.0728 fingerprint, the source case haplotype was circulating among endemic cases prior to the outbreak. Genetic and temporal distances were not correlated for either RFLP 0.0142 (r2 = - 0.05) or RFLP 0.0728 (r2 = 0.09) when all isolates were analyzed. CONCLUSIONS: We found no evidence that endemic strains acquired mutations resulting in their emergence in outbreak form. We conclude that the propagation of these outbreaks was likely driven by the combination of characteristics of the source cases, contacts, and the environment. The role of whole-genome sequencing in understanding mycobacterial evolution and in assisting public health authorities in conducting contact investigations and managing outbreaks is important and expected to grow in the future.

Adaptation in a Fibronectin Binding Autolysin of Staphylococcus saprophyticus.

Human-pathogenic bacteria are found in a variety of niches, including free-living, zoonotic, and microbiome environments. Identifying bacterial adaptations that enable invasive disease is an important means of gaining insight into the molecular basis of pathogenesis and understanding pathogen emergence. Staphylococcus saprophyticus, a leading cause of urinary tract infections, can be found in the environment, food, animals, and the human microbiome. We identified a selective sweep in the gene encoding the Aas adhesin, a key virulence factor that binds host fibronectin. We hypothesize that the mutation under selection (aas_2206A>C) facilitates colonization of the urinary tract, an environment where bacteria are subject to strong shearing forces. The mutation appears to have enabled emergence and expansion of a human-pathogenic lineage of S. saprophyticus. These results demonstrate the power of evolutionary genomic approaches in discovering the genetic basis of virulence and emphasize the pleiotropy and adaptability of bacteria occupying diverse niches. IMPORTANCEStaphylococcus saprophyticus is an important cause of urinary tract infections (UTI) in women; such UTI are common, can be severe, and are associated with significant impacts to public health. In addition to being a cause of human UTI, S. saprophyticus can be found in the environment, in food, and associated with animals. After discovering that UTI strains of S. saprophyticus are for the most part closely related to each other, we sought to determine whether these strains are specially adapted to cause disease in humans. We found evidence suggesting that a mutation in the gene aas is advantageous in the context of human infection. We hypothesize that the mutation allows S. saprophyticus to survive better in the human urinary tract. These results show how bacteria found in the environment can evolve to cause disease.