Effects of urban-induced mutations on ecology, evolution and health.

Increasing evidence suggests that urbanization is associated with higher mutation rates, which can affect the health and evolution of organisms that inhabit cities. Elevated pollution levels in urban areas can induce DNA damage, leading to de novo mutations. Studies on mutations induced by urban pollution are most prevalent in humans and microorganisms, whereas studies of non-human eukaryotes are rare, even though increased mutation rates have the potential to affect organisms and their populations in contemporary time. Our Perspective explores how higher mutation rates in urban environments could impact the fitness, ecology and evolution of populations. Most mutations will be neutral or deleterious, and higher mutation rates associated with elevated pollution in urban populations can increase the risk of cancer in humans and potentially other species. We highlight the potential for urban-driven increased deleterious mutational loads in some organisms, which could lead to a decline in population growth of a wide diversity of organisms. Although beneficial mutations are expected to be rare, we argue that higher mutation rates in urban areas could influence adaptive evolution, especially in organisms with short generation times. Finally, we explore avenues for future research to better understand the effects of urban-induced mutations on the fitness, ecology and evolution of city-dwelling organisms.

Moving psychiatric deinstitutionalization forward: A scoping review of barriers and facilitators.

Psychiatric deinstitutionalization (PDI) processes aim to transform long-term psychiatric care by closing or reducing psychiatric hospitals, reallocating beds, and establishing comprehensive community-based services for individuals with severe and persistent mental health difficulties. This scoping review explores the extensive literature on PDI, spanning decades, regions, socio-political contexts, and disciplines, to identify barriers and facilitators of PDI implementation, providing researchers and policymakers with a categorization of these factors. To identify barriers and facilitators, three electronic databases (Medline, CINAHL, and Sociological Abstracts) were searched, yielding 2,250 references. After screening and reviewing, 52 studies were included in the final analysis. Thematic synthesis was utilized to categorize the identified factors, responding to the review question. The analysis revealed that barriers to PDI include inadequate planning, funding, and leadership, limited knowledge, competing interests, insufficient community-based alternatives, and resistance from the workforce, community, and family/caregivers. In contrast, facilitators encompass careful planning, financing and coordination, available research and evidence, strong and sustained advocacy, comprehensive community services, and a well-trained workforce engaged in the process. Exogenous factors, such as conflict and humanitarian disasters, can also play a role in PDI processes. Implementing PDI requires a multifaceted strategy, strong leadership, diverse stakeholder participation, and long-term political and financial support. Understanding local needs and forces is crucial, and studying PDI necessitates methodological flexibility and sensitivity to contextual variation. At the same time, based on the development of the review itself, we identify four limitations in the literature, concerning "time," "location," "focus," and "voice." We call for a renewed research and advocacy agenda around this neglected aspect of contemporary global mental health policy is needed.

Transposons contribute to the functional diversification of the head, gut, and ovary transcriptomes across Drosophila natural strains.

Transcriptomes are dynamic, with cells, tissues, and body parts expressing particular sets of transcripts. Transposable elements (TEs) are a known source of transcriptome diversity; however, studies often focus on a particular type of chimeric transcript, analyze single body parts or cell types, or are based on incomplete TE annotations from a single reference genome. In this work, we have implemented a method based on de novo transcriptome assembly that minimizes the potential sources of errors while identifying a comprehensive set of gene-TE chimeras. We applied this method to the head, gut, and ovary dissected from five Drosophila melanogaster natural strains, with individual reference genomes available. We found that ∼19% of body part-specific transcripts are gene-TE chimeras. Overall, chimeric transcripts contribute a mean of 43% to the total gene expression, and they provide protein domains for DNA binding, catalytic activity, and DNA polymerase activity. Our comprehensive data set is a rich resource for follow-up analysis. Moreover, because TEs are present in virtually all species sequenced to date, their role in spatially restricted transcript expression is likely not exclusive to the species analyzed in this work.

Two-step CRISPR-Cas9 protocol for transposable element deletion in D. melanogaster natural populations.

We present a protocol for generating a precise deletion, without altering the genetic background of the strain, of a transposable element (TE) in a natural population of Drosophila melanogaster using two steps of CRISPR-Cas9 homology-directed repair. We describe steps for replacing the TE by a fluorescent marker and for subsequent marker removal using single-guide RNAs, repair plasmids, and microinjection. We also detail steps for screening the deletion of the TE and generating a homozygous mutant strain. For complete details on the use and execution of this protocol, please refer to Merenciano and Gonzalez.1.

DrosOmics: A Browser to Explore -omics Variation Across High-Quality Reference Genomes From Natural Populations of Drosophila melanogaster.

The advent of long-read sequencing technologies has allowed the generation of multiple high-quality de novo genome assemblies for multiple species, including well-known model species such as Drosophila melanogaster. Genome assemblies for multiple individuals of the same species are key to discover the genetic diversity present in natural populations, especially the one generated by transposable elements, the most common type of structural variant. Despite the availability of multiple genomic data sets for D. melanogaster populations, we lack an efficient visual tool to display different genome assemblies simultaneously. In this work, we present DrosOmics, a population genomic-oriented browser currently containing 52 high-quality reference genomes of D. melanogaster, including annotations from a highly reliable set of transposable elements, and functional transcriptomics and epigenomics data for 26 genomes. DrosOmics is based on JBrowse 2, a highly scalable platform, which allows the visualization of multiple assemblies at once, key to unraveling structural and functional features of D. melanogaster natural populations. DrosOmics is an open access browser and is freely available at http://gonzalezlab.eu/drosomics.

Gene expression differences consistent with water loss reduction underlie desiccation tolerance of natural Drosophila populations.

BACKGROUND: Climate change is one of the main factors shaping the distribution and biodiversity of organisms, among others by greatly altering water availability, thus exposing species and ecosystems to harsh desiccation conditions. However, most of the studies so far have focused on the effects of increased temperature. Integrating transcriptomics and physiology is key to advancing our knowledge on how species cope with desiccation stress, and these studies are still best accomplished in model organisms. RESULTS: Here, we characterized the natural variation of European D. melanogaster populations across climate zones and found that strains from arid regions were similar or more tolerant to desiccation compared with strains from temperate regions. Tolerant and sensitive strains differed not only in their transcriptomic response to stress but also in their basal expression levels. We further showed that gene expression changes in tolerant strains correlated with their physiological response to desiccation stress and with their cuticular hydrocarbon composition, and functionally validated three of the candidate genes identified. Transposable elements, which are known to influence stress response across organisms, were not found to be enriched nearby differentially expressed genes. Finally, we identified several tRNA-derived small RNA fragments that differentially targeted genes in response to desiccation stress. CONCLUSIONS: Overall, our results showed that basal gene expression differences across individuals should be analyzed if we are to understand the genetic basis of differential stress survival. Moreover, tRNA-derived small RNA fragments appear to be relevant across stress responses and allow for the identification of stress-response genes not detected at the transcriptional level.

The Interplay Between Developmental Stage and Environment Underlies the Adaptive Effect of a Natural Transposable Element Insertion.

Establishing causal links between adaptive mutations and ecologically relevant phenotypes is key to understanding the process of adaptation, which is a central goal in evolutionary biology with applications for conservation, medicine, and agriculture. Yet despite recent progress, the number of identified causal adaptive mutations remains limited. Linking genetic variation to fitness-related effects is complicated by gene-by-gene and gene-by-environment interactions, among other processes. Transposable elements, which are often ignored in the quest for the genetic basis of adaptive evolution, are a genome-wide source of regulatory elements across organisms that can potentially result in adaptive phenotypes. In this work, we combine gene expression, in vivo reporter assays, CRISPR/Cas9 genome editing, and survival experiments to characterize in detail the molecular and phenotypic consequences of a natural Drosophila melanogaster transposable element insertion: the roo solo-LTR FBti0019985. This transposable element provides an alternative promoter to the transcription factor Lime, involved in cold- and immune-stress responses. We found that the effect of FBti0019985 on Lime expression depends on the interplay between the developmental stage and environmental condition. We further establish a causal link between the presence of FBti0019985 and increased survival to cold- and immune-stress. Our results exemplify how several developmental stages and environmental conditions need to be considered to characterize the molecular and functional effects of a genetic variant, and add to the growing body of evidence that transposable elements can induce complex mutations with ecologically relevant effects.

Imaging in atrial fibrillation: A way to assess atrial fibrosis and remodeling to assist decision-making.

The 2020 ESC atrial fibrillation (AF) guidelines suggest the novel 4S-AF scheme for the characterization of AF. Imaging techniques could be helpful for this objective in everyday clinical practice, and information derived from these techniques reflects basic aspects of the pathophysiology of AF, which may facilitate treatment decision-making, and optimal management of AF patients. The aim of this review is to provide an overview of the mechanisms associated with atrial fibrosis and to describe imaging techniques that may help the management of AF patients in clinical practice. Transthoracic echocardiography is the most common procedure given its versatility, safety, and simplicity. Transesophageal echocardiography provides higher resolution exploration, and speckle tracking echocardiography can provide incremental functional and prognostic information over conventional echocardiographic parameters. In addition, LA deformation imaging, including LA strain and strain rate, are related to the extent of fibrosis. On the other hand, multidetector-row computed tomography and cardiac magnetic resonance provide higher resolution data and more accurate assessment of the dimensions, structure, and spatial relationships of the LA. Imaging is central when deciding on catheter ablation or cardioversion, and helps in selecting those patients who will really benefit from these procedures. Moreover, imaging enhances the understanding of the underlying mechanisms of atrial remodeling and might assists in refining the risk of stroke, which help to select the best medical therapies/interventions. In summary, evaluation of LA enlargement, LA remodeling and fibrosis with imaging techniques adds clinical and prognostic information and should be assessed as a part of routine comprehensive AF evaluation.

Experimental Validation of Transposable Element Insertions Using the Polymerase Chain Reaction (PCR).

Transposable elements (TEs), also known as transposons, are repetitive DNA sequences, present in virtually all organisms, that can move from one genomic position to another. TEs can be a source of mutations with important consequences for organisms. Despite their interest, its repetitive nature has made their study very challenging. However, the emergence of new sequencing technologies that allow obtaining long-read sequences, has improved the in silico de novo detection and annotation of TEs. The de novo annotation of TEs has already been performed in several organisms including the fruit fly Drosophila melanogaster. Yet, experimental validation can be used to confirm the presence of TEs in specific D. melanogaster natural populations. Here, we present a step-by-step protocol to experimentally validate by polymerase chain reaction (PCR) the presence and/or absence of TEs in natural populations of D. melanogaster. This detailed protocol has been implemented in the participant high schools of the Citizen Fly Lab activity that is part of the international citizen science project Melanogaster: Catch the Fly! ( https://melanogaster.eu ). Specifically, the students collaborate with the scientists of the European Drosophila Population Genomics Consortium (DrosEU) in the experimental validation of new genetic variants, previously identified using bioinformatic techniques.

The genomic basis of copper tolerance in Drosophila is shaped by a complex interplay of regulatory and environmental factors.

BACKGROUND: Escalation in industrialization and anthropogenic activity have resulted in an increase of pollutants released into the environment. Of these pollutants, heavy metals such as copper are particularly concerning due to their bio-accumulative nature. Due to its highly heterogeneous distribution and its dual nature as an essential micronutrient and toxic element, the genetic basis of copper tolerance is likely shaped by a complex interplay of genetic and environmental factors. RESULTS: In this study, we utilized the natural variation present in multiple populations of Drosophila melanogaster collected across Europe to screen for variation in copper tolerance. We found that latitude and the degree of urbanization at the collection sites, rather than any other combination of environmental factors, were linked to copper tolerance. While previously identified copper-related genes were not differentially expressed in tolerant vs. sensitive strains, genes involved in metabolism, reproduction, and protease induction contributed to the differential stress response. Additionally, the greatest transcriptomic and physiological responses to copper toxicity were seen in the midgut, where we found that preservation of gut acidity is strongly linked to greater tolerance. Finally, we identified transposable element insertions likely to play a role in copper stress response. CONCLUSIONS: Overall, by combining genome-wide approaches with environmental association analysis, and functional analysis of candidate genes, our study provides a unique perspective on the genetic and environmental factors that shape copper tolerance in natural D. melanogaster populations and identifies new genes, transposable elements, and physiological traits involved in this complex phenotype.

Promoting scientific literacy in evolution through citizen science.

Evolutionary understanding is central to biology. It is also an essential prerequisite to understanding and making informed decisions about societal issues such as climate change. Yet, evolution is generally poorly understood by civil society and many misconceptions exist. Citizen science, which has been increasing in popularity as a means to gather new data and promote scientific literacy, is one strategy through which people could learn about evolution. However, despite the potential for citizen science to promote evolution learning opportunities, very few projects implement them. In this paper, we make the case for incorporating evolution education into citizen science, define key learning goals, and suggest opportunities for designing and evaluating projects in order to promote scientific literacy in evolution.

Population-scale long-read sequencing uncovers transposable elements associated with gene expression variation and adaptive signatures in Drosophila.

High quality reference genomes are crucial to understanding genome function, structure and evolution. The availability of reference genomes has allowed us to start inferring the role of genetic variation in biology, disease, and biodiversity conservation. However, analyses across organisms demonstrate that a single reference genome is not enough to capture the global genetic diversity present in populations. In this work, we generate 32 high-quality reference genomes for the well-known model species D. melanogaster and focus on the identification and analysis of transposable element variation as they are the most common type of structural variant. We show that integrating the genetic variation across natural populations from five climatic regions increases the number of detected insertions by 58%. Moreover, 26% to 57% of the insertions identified using long-reads were missed by short-reads methods. We also identify hundreds of transposable elements associated with gene expression variation and new TE variants likely to contribute to adaptive evolution in this species. Our results highlight the importance of incorporating the genetic variation present in natural populations to genomic studies, which is essential if we are to understand how genomes function and evolve.

Transposable element variants and their potential adaptive impact in urban populations of the malaria vector Anopheles coluzzii.

Anopheles coluzzii is one of the primary vectors of human malaria in sub-Saharan Africa. Recently, it has spread into the main cities of Central Africa threatening vector control programs. The adaptation of An. coluzzii to urban environments partly results from an increased tolerance to organic pollution and insecticides. Some of the molecular mechanisms for ecological adaptation are known, but the role of transposable elements (TEs) in the adaptive processes of this species has not been studied yet. As a first step toward assessing the role of TEs in rapid urban adaptation, we sequenced using long reads six An. coluzzii genomes from natural breeding sites in two major Central Africa cities. We de novo annotated TEs in these genomes and in an additional high-quality An. coluzzii genome, and we identified 64 new TE families. TEs were nonrandomly distributed throughout the genome with significant differences in the number of insertions of several superfamilies across the studied genomes. We identified seven putatively active families with insertions near genes with functions related to vectorial capacity, and several TEs that may provide promoter and transcription factor binding sites to insecticide resistance and immune-related genes. Overall, the analysis of multiple high-quality genomes allowed us to generate the most comprehensive TE annotation in this species to date and identify several TE insertions that could potentially impact both genome architecture and the regulation of functionally relevant genes. These results provide a basis for future studies of the impact of TEs on the biology of An. coluzzii.

Regulatory regions in natural transposable element insertions drive interindividual differences in response to immune challenges in Drosophila.

BACKGROUND: Variation in gene expression underlies interindividual variability in relevant traits including immune response. However, the genetic variation responsible for these gene expression changes remains largely unknown. Among the non-coding variants that could be relevant, transposable element insertions are promising candidates as they have been shown to be a rich and diverse source of cis-regulatory elements. RESULTS: In this work, we use a population genetics approach to identify transposable element insertions likely to increase the tolerance of Drosophila melanogaster to bacterial infection by affecting the expression of immune-related genes. We identify 12 insertions associated with allele-specific expression changes in immune-related genes. We experimentally validate three of these insertions including one likely to be acting as a silencer, one as an enhancer, and one with a dual role as enhancer and promoter. The direction in the change of gene expression associated with the presence of several of these insertions is consistent with an increased survival to infection. Indeed, for one of the insertions, we show that this is the case by analyzing both natural populations and CRISPR/Cas9 mutants in which the insertion is deleted from its native genomic context. CONCLUSIONS: We show that transposable elements contribute to gene expression variation in response to infection in D. melanogaster and that this variation is likely to affect their survival capacity. Because the role of transposable elements as regulatory elements is not restricted to Drosophila, transposable elements are likely to play a role in immune response in other organisms as well.

The discovery, distribution, and diversity of DNA viruses associated with Drosophila melanogaster in Europe.

Drosophila melanogaster is an important model for antiviral immunity in arthropods, but very few DNA viruses have been described from the family Drosophilidae. This deficiency limits our opportunity to use natural host-pathogen combinations in experimental studies, and may bias our understanding of the Drosophila virome. Here, we report fourteen DNA viruses detected in a metagenomic analysis of 6668 pool-sequenced Drosophila, sampled from forty-seven European locations between 2014 and 2016. These include three new nudiviruses, a new and divergent entomopoxvirus, a virus related to Leptopilina boulardi filamentous virus, and a virus related to Musca domestica salivary gland hypertrophy virus. We also find an endogenous genomic copy of galbut virus, a double-stranded RNA partitivirus, segregating at very low frequency. Remarkably, we find that Drosophila Vesanto virus, a small DNA virus previously described as a bidnavirus, may be composed of up to twelve segments and thus represent a new lineage of segmented DNA viruses. Two of the DNA viruses, Drosophila Kallithea nudivirus and Drosophila Vesanto virus are relatively common, found in 2 per cent or more of wild flies. The others are rare, with many likely to be represented by a single infected fly. We find that virus prevalence in Europe reflects the prevalence seen in publicly available datasets, with Drosophila Kallithea nudivirus and Drosophila Vesanto virus the only ones commonly detectable in public data from wild-caught flies and large population cages, and the other viruses being rare or absent. These analyses suggest that DNA viruses are at lower prevalence than RNA viruses in D.melanogaster, and may be less likely to persist in laboratory cultures. Our findings go some way to redressing an earlier bias toward RNA virus studies in Drosophila, and lay the foundation needed to harness the power of Drosophila as a model system for the study of DNA viruses.

Broad geographic sampling reveals the shared basis and environmental correlates of seasonal adaptation in Drosophila.

To advance our understanding of adaptation to temporally varying selection pressures, we identified signatures of seasonal adaptation occurring in parallel among Drosophila melanogaster populations. Specifically, we estimated allele frequencies genome-wide from flies sampled early and late in the growing season from 20 widely dispersed populations. We identified parallel seasonal allele frequency shifts across North America and Europe, demonstrating that seasonal adaptation is a general phenomenon of temperate fly populations. Seasonally fluctuating polymorphisms are enriched in large chromosomal inversions, and we find a broad concordance between seasonal and spatial allele frequency change. The direction of allele frequency change at seasonally variable polymorphisms can be predicted by weather conditions in the weeks prior to sampling, linking the environment and the genomic response to selection. Our results suggest that fluctuating selection is an important evolutionary force affecting patterns of genetic variation in Drosophila.

Benchmarking the performance of Pool-seq SNP callers using simulated and real sequencing data.

Population genomics is a fast-developing discipline with promising applications in a growing number of life sciences fields. Advances in sequencing technologies and bioinformatics tools allow population genomics to exploit genome-wide information to identify the molecular variants underlying traits of interest and the evolutionary forces that modulate these variants through space and time. However, the cost of genomic analyses of multiple populations is still too high to address them through individual genome sequencing. Pooling individuals for sequencing can be a more effective strategy in Single Nucleotide Polymorphism (SNP) detection and allele frequency estimation because of a higher total coverage. However, compared to individual sequencing, SNP calling from pools has the additional difficulty of distinguishing rare variants from sequencing errors, which is often avoided by establishing a minimum threshold allele frequency for the analysis. Finding an optimal balance between minimizing information loss and reducing sequencing costs is essential to ensure the success of population genomics studies. Here, we have benchmarked the performance of SNP callers for Pool-seq data, based on different approaches, under different conditions, and using computer simulations and real data. We found that SNP callers performance varied for allele frequencies up to 0.35. We also found that SNP callers based on Bayesian (SNAPE-pooled) or maximum likelihood (MAPGD) approaches outperform the two heuristic callers tested (VarScan and PoolSNP), in terms of the balance between sensitivity and FDR both in simulated and sequencing data. Our results will help inform the selection of the most appropriate SNP caller not only for large-scale population studies but also in cases where the Pool-seq strategy is the only option, such as in metagenomic or polyploid studies.

Temperature, rainfall and wind variables underlie environmental adaptation in natural populations of Drosophila melanogaster.

While several studies in a diverse set of species have shed light on the genes underlying adaptation, our knowledge on the selective pressures that explain the observed patterns lags behind. Drosophila melanogaster is a valuable organism to study environmental adaptation because this species originated in Southern Africa and has recently expanded worldwide, and also because it has a functionally well-annotated genome. In this study, we aimed to decipher which environmental variables are relevant for adaptation of D. melanogaster natural populations in Europe and North America. We analysed 36 whole-genome pool-seq samples of D. melanogaster natural populations collected in 20 European and 11 North American locations. We used the BayPass software to identify single nucleotide polymorphisms (SNPs) and transposable elements (TEs) showing signature of adaptive differentiation across populations, as well as significant associations with 59 environmental variables related to temperature, rainfall, evaporation, solar radiation, wind, daylight hours, and soil type. We found that in addition to temperature and rainfall, wind related variables are also relevant for D. melanogaster environmental adaptation. Interestingly, 23%-51% of the genes that showed significant associations with environmental variables were not found overly differentiated across populations. In addition to SNPs, we also identified 10 reference transposable element insertions associated with environmental variables. Our results showed that genome-environment association analysis can identify adaptive genetic variants that are undetected by population differentiation analysis while also allowing the identification of candidate environmental drivers of adaptation.

Identifying chromosomal subpopulations based on their recombination histories advances the study of the genetic basis of phenotypic traits.

Recombination is a main source of genetic variability. However, the potential role of the variation generated by recombination in phenotypic traits, including diseases, remains unexplored because there is currently no method to infer chromosomal subpopulations based on recombination pattern differences. We developed recombClust, a method that uses SNP-phased data to detect differences in historic recombination in a chromosome population. We validated our method by performing simulations and by using real data to accurately predict the alleles of well-known recombination modifiers, including common inversions in Drosophila melanogaster and human, and the chromosomes under selective pressure at the lactase locus in humans. We then applied recombClust to the complex human 1q21.1 region, where nonallelic homologous recombination produces deleterious phenotypes. We discovered and validated the presence of two different recombination histories in these regions that significantly associated with the differential expression of ANKRD35 in whole blood and that were in high linkage with variants previously associated with hypertension. By detecting differences in historic recombination, our method opens a way to assess the influence of recombination variation in phenotypic traits.