ABSTRACT Invasive rodents cause severe ecosystem degradation on islands and can be challenging to eradicate. Current best‐practices rely on the application of toxic oral baits and have led to successful eradications and remarkable recoveries of native flora and fauna. Yet this single method is not universally applicable. Genetic biocontrol offers a suite of new solutions to potentially improve outcomes in the critical “last mile” of eradication. These approaches involve the release of genetically modified individuals of the target species to reduce population fitness over time. These include self‐limiting approaches which require multiple releases and self‐sustaining mechanisms (i.e., select gene‐drive systems) which could theoretically collapse populations after a single release. While gene drive systems have received significant attention, their development in vertebrates remains technically challenging, and their ecological and regulatory implications are still in active debate. In contrast, some non‐drive genetic biocontrol approaches, such as Y‐linked editors, fsRIDL, and Gravid Lethal, offer self‐limiting alternatives that may be more immediately deployable. These approaches could be used to supplement toxicant‐based methods and may also have reduced environmental risks and regulatory barriers. To evaluate the potential of these tools, we developed an individual‐based model simulating the eradication of house mice (Mus musculus) on a 125 ha island with an initial population of 11,000 individuals. We tested various combinations of genetic biocontrol release and effort strategies to understand tradeoffs between required effort and uncertainty for achieving successful eradication. Under certain effort strategies, we found that the Gravid Lethal approach performed the best, achieving eradication in < 2.3 years with intensive release effort and monitoring. Our findings suggest that integrating non‐drive genetic biocontrol into adaptive management frameworks could enhance the effectiveness and feasibility of rodent eradication programs. These tools are not replacements for toxicants but may serve as critical supplements—particularly in the “last mile” of eradication.

In this study, we used a multi-faceted approach to understand patterns of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission and persistence in a wild white-tailed deer (Odocoileus virginianus) population. Serology data indicated transmission of SARS-CoV-2 and persistence during the seven-month sampling period. Traditional disease modelling based on deer-to-deer transmission indicated relatively low prevalence with an R0 of 1.9 and recovery period of 7 days; however, individual-based modelling informed by GPS tracked-movement data captured a potential transmission event. Phylogenetic analyses revealed a recurring pattern of divergent groups of deer-derived sequences with human-derived sequences falling close to each deer-derived cluster. Further, human-derived sequences were frequently sampled months prior to the deer-derived sequences, indicating repeated human to deer spillover. Using multiple types of data as well as both fine and broad scale analyses, we have characterized a pattern of localized outbreaks of SARS-CoV-2 within white-tailed deer populations that are likely recurring due to frequent spillover events. Our results suggest that while deer-to-deer transmission occurs over small spatiotemporal scales, SARS-CoV-2 persistence over longer periods and across larger regions is likely driven by repeated spillover from human populations.

Cities present unique ecological challenges and opportunities for species that have adapted to human-dominated environments. To understand how urban landscape heterogeneity shapes the movements of an iconic urban animal, the brown rat ( Rattus norvegicus ), we investigated the factors influencing rat genetic connectivity across Manhattan, New York City. We generated genome-wide SNP data from rats sampled throughout Manhattan as well as paired surface-subway locations, created a habitat suitability model using systematic municipal rat survey data, and employed landscape genetic approaches to identify urban features influencing rat gene flow. Our models revealed that underground subway tunnels were the strongest facilitator of rat gene flow, with municipal zoning also strongly influencing connectivity, particularly through low / medium-density residential areas. Habitat suitability analysis showed that high human population density, old building ages, and lower median household income were the strongest predictors of rat presence. Subterranean sampling revealed that rats in subway stations exhibited greater genetic relatedness than surface rats at short distances but lower inbreeding coefficients, suggesting underground tunnels facilitate longer-distance dispersal while maintaining some connectivity to surface populations. These findings demonstrate that physical infrastructure, political, and socioeconomic factors shape urban rat ecology, with implications for more targeted approaches to pest management in complex urban environments. This study is the first comprehensive analysis of rat dispersal and genetic connectivity in relation to complex cityscape heterogeneity. ### Competing Interest Statement The authors have declared no competing interest. National Science FoundationNational Science Foundation, , DEB-1457523, DBI-1531639

Tolerance and resistance are two host eco-immunological strategies in response to microparasite invasion. In the strategy of “resistance”, host responses are induced to decrease microparasite replication while the “tolerance” strategy allows hosts coexistence with microparasites by minimizing responses to avoid immune-mediated damage. The causative agent of Lyme disease is a group of genotypically diverse bacterial species, Borrelia burgdorferi sensu lato (Bb), which is transmitted by Ixodes ticks and persists in different reservoir animals. In North America, eastern fence lizards (Sceloporus undulatus) can be fed on by Ixodes ticks but are incompetent to one genotype of Bb (i.e., ospC type A). However, field-collected lizards showed evidence of previous infection by Bb strains with undefined genotypes. Supporting this evidence, we introduced three genotypically different Bb strains individually to eastern fence lizards and found a Bb genotype-dependent manner of infectivity. We compared liver transcriptomics and observed elevated immune responses triggered by a lizard-incompetent Bb strain (strain B31). We showed two lizard-competent strains with one having no immunomodulation (strain B379) but the other developing upregulated immune responses (strain 297). These results suggest that genetic variation in microparasites both induces different host strategies for dealing with infection and determines microparasite fitness in the hosts. These findings demonstrate that Bb and eastern fence lizards can serve as a model to investigate the mechanisms underlying eco-immunological strategies of tolerance vs. resistance during host-microparasite interaction.

Engineered gene drives create potential for both widespread benefits and irreversible harms to ecosystems. CRISPR-based systems of allelic conversion have rapidly accelerated gene drive research across diverse taxa, putting field trials and their necessary risk assessments on the horizon. Dynamic process-based models provide flexible quantitative platforms to predict gene drive outcomes in the context of system-specific ecological and evolutionary features. Here, we synthesize gene drive dynamic modeling studies to highlight research trends, knowledge gaps, and emergent principles, organized around their genetic, demographic, spatial, environmental, and implementation features. We identify the phenomena that most significantly influence model predictions, discuss limitations of biological complexity and uncertainty, and provide insights to promote responsible development and model-assisted risk assessment of gene drives.

The range of hosts a pathogen can infect is a key trait, influencing human disease risk and reservoir host infection dynamics. Borrelia burgdorferi sensu stricto (Bb), an emerging zoonotic pathogen, causes Lyme disease and is widely considered a host generalist, commonly infecting mammals and birds. Yet the extent of intraspecific variation in Bb host breadth, its role in determining host competence, and potential implications for human infection remain unclear. We conducted a long-term study of Bb diversity, defined by the polymorphic ospC locus, across white-footed mice, passerine birds, and tick vectors, leveraging long-read amplicon sequencing. Our results reveal strong variation in host breadth across Bb genotypes, exposing a spectrum of genotype-specific host-adapted phenotypes. We found support for multiple niche polymorphism, maintaining Bb diversity in nature and little evidence of temporal shifts in genotype dominance, as would be expected under negative frequency-dependent selection. Passerine birds support the circulation of several human-invasive strains (HISs) in the local tick population and harbor greater Bb genotypic diversity compared with white-footed mice. Mouse-adapted Bb genotypes exhibited longer persistence in individual mice compared with nonadapted genotypes. Genotype communities infecting individual mice preferentially became dominated by mouse-adapted genotypes over time. We posit that intraspecific variation in Bb host breadth and adaptation helps maintain overall species fitness in response to transmission by a generalist vector.

The ongoing COVID-19 pandemic is a stark reminder of the devastating consequences of pathogen spillover from wildlife to human hosts, particularly in densely populated urban centers. Prevention of future zoonotic disease is contingent on informed surveillance for known and novel threats across diverse human-wildlife interfaces. Cities are a key venue for potential spillover events because of the presence of zoonotic pathogens transmitted by hosts and vectors living in close proximity to dense human settlements. Effectively identifying and managing zoonotic hazards requires understanding the socio-ecological processes driving hazard distribution and pathogen prevalence in dynamic and heterogeneous urban landscapes. Despite increasing awareness of the human health impacts of zoonotic hazards, the integration of an eco-epidemiological perspective into public health management plans remains limited. Here we discuss how landscape patterns, abiotic conditions, and biotic interactions influence zoonotic hazards across highly urbanized cities (HUCs) in temperate climates to promote their efficient and effective management by a multi-sectoral coalition of public health stakeholders. We describe how to interpret both direct and indirect ecological processes, incorporate spatial scale, and evaluate networks of connectivity specific to different zoonotic hazards to promote biologically-informed and targeted decision-making. Using New York City, USA as a case study, we identify major zoonotic threats, apply knowledge of relevant ecological factors, and highlight opportunities and challenges for research and intervention. We aim to broaden the toolbox of urban public health stakeholders by providing ecologically-informed, practical guidance for the evaluation and management of zoonotic hazards.

Predicting pathogen emergence and spillover risk requires understanding the determinants of a pathogens' host range and the traits involved in host competence. While host competence is often considered a fixed species-specific trait, it may be variable if pathogens diversify across hosts. Balancing selection can lead to maintenance of pathogen polymorphisms (multiple-niche-polymorphism; MNP). The causative agent of Lyme disease, Borrelia burgdorferi (Bb), provides a model to study the evolution of host adaptation, as some Bb strains defined by their outer surface protein C (ospC) genotype, are widespread in white-footed mice and others are associated with non-rodent vertebrates (e.g. birds). To identify the mechanisms underlying potential strain × host adaptation, we infected American robins and white-footed mice, with three Bb strains of different ospC genotypes. Bb burdens varied by strain in a host-dependent fashion, and strain persistence in hosts largely corresponded to Bb survival at early infection stages and with transmission to larvae (i.e. fitness). Early survival phenotypes are associated with cell adhesion, complement evasion and/or inflammatory and antibody-mediated removal of Bb, suggesting directional selective pressure for host adaptation and the potential role of MNP in maintaining OspC diversity. Our findings will guide future investigations to inform eco-evolutionary models of host adaptation for microparasites.

Host association-the selective adaptation of pathogens to specific host species-evolves through constant interactions between host and pathogens, leaving a lot yet to be discovered on immunological mechanisms and genomic determinants. The causative agents of Lyme disease (LD) are spirochete bacteria composed of multiple species of the Borrelia burgdorferi sensu lato complex, including B. burgdorferi (Bb), the main LD pathogen in North America-a useful model for the study of mechanisms underlying host-pathogen association. Host adaptation requires pathogens' ability to evade host immune responses, such as complement, the first-line innate immune defense mechanism. We tested the hypothesis that different host-adapted phenotypes among Bb strains are linked to polymorphic loci that confer complement evasion traits in a host-specific manner. We first examined the survivability of 20 Bb strains in sera in vitro and/or bloodstream and tissues in vivo from rodent and avian LD models. Three groups of complement-dependent host-association phenotypes emerged. We analyzed complement-evasion genes, identified a priori among all strains and sequenced and compared genomes for individual strains representing each phenotype. The evolutionary history of ospC loci is correlated with host-specific complement-evasion phenotypes, while comparative genomics suggests that several gene families and loci are potentially involved in host association. This multidisciplinary work provides novel insights into the functional evolution of host-adapted phenotypes, building a foundation for further investigation of the immunological and genomic determinants of host association. IMPORTANCE Host association is the phenotype that is commonly found in many pathogens that preferential survive in particular hosts. The Lyme disease (LD)-causing agent, B. burgdorferi (Bb), is an ideal model to study host association, as Bb is mainly maintained in nature through rodent and avian hosts. A widespread yet untested concept posits that host association in Bb strains is linked to Bb functional genetic variation conferring evasion to complement, an innate defense mechanism in vertebrate sera. Here, we tested this concept by grouping 20 Bb strains into three complement-dependent host-association phenotypes based on their survivability in sera and/or bloodstream and distal tissues in rodent and avian LD models. Phylogenomic analysis of these strains further correlated several gene families and loci, including ospC, with host-specific complement-evasion phenotypes. Such multifaceted studies thus pave the road to further identify the determinants of host association, providing mechanistic insights into host-pathogen interaction.

Brown rats (Rattus norvegicus) thrive in urban environments by navigating the anthropocentric environment and taking advantage of human resources and by-products. From the human perspective, rats are a chronic problem that causes billions of dollars in damage to agriculture, health, and infrastructure. Did genetic adaptation play a role in the spread of rats in cities? To approach this question, we collected whole-genome sequences from 29 brown rats from New York City (NYC) and scanned for genetic signatures of adaptation. We tested for 1) high-frequency, extended haplotypes that could indicate selective sweeps and 2) loci of extreme genetic differentiation between the NYC sample and a sample from the presumed ancestral range of brown rats in northeast China. We found candidate selective sweeps near or inside genes associated with metabolism, diet, the nervous system, and locomotory behavior. Patterns of differentiation between NYC and Chinese rats at putative sweep loci suggest that many sweeps began after the split from the ancestral population. Together, our results suggest several hypotheses on adaptation in rats living in proximity to humans.

Urban Norway rats ( Rattus norvegicus ) carry several pathogens transmissible to people. However, pathogen prevalence can vary across fine spatial scales (i.e., by city block). Using a population genomics approach, we sought to describe rat movement patterns across an urban landscape and to evaluate whether these patterns align with pathogen distributions. We genotyped 605 rats from a single neighborhood in Vancouver, Canada, and used 1,495 genome‐wide single nucleotide polymorphisms to identify parent–offspring and sibling relationships using pedigree analysis. We resolved 1,246 pairs of relatives, of which only 1% of pairs were captured in different city blocks. Relatives were primarily caught within 33 meters of each other leading to a highly leptokurtic distribution of dispersal distances. Using binomial generalized linear mixed models, we evaluated whether family relationships influenced rat pathogen status with the bacterial pathogens Leptospira interrogans , Bartonella tribocorum , and Clostridium difficile , and found that an individual's pathogen status was not predicted any better by including disease status of related rats. The spatial clustering of related rats and their pathogens lends support to the hypothesis that spatially restricted movement promotes the heterogeneous patterns of pathogen prevalence evidenced in this population. Our findings also highlight the utility of evolutionary tools to understand movement and rat‐associated health risks in urban landscapes.

ABSTRACTHost specialization is an ecological and evolutionary process by which a pathogen becomes differentially adapted to a subset of hosts, restricting its host range. For parasites transmitted by generalist vectors, host specialization is not expected to evolve because of the decreased survival of those parasites in inadequate hosts. Thus, parasites may develop adaptation strategies, resulting in host specialization. The causative agents of Lyme disease are multiple species of bacteria,Borrelia burgdorferisensu lato species complex (Bbsl), and are suitable for examining host specialization as birds and rodents were found to carry different species of these bacteria. Debate exists on whether host specialization occurs among these strains within a particular species ofBbsl, such asB. burgdorferisensu stricto (Bbss). Current evidence supports someBbss strains are widespread in white-footed mice but others are in non-rodent vertebrates, such as birds. To recapitulate specialization in the laboratory and define the mechanisms for host specialization, we introduced different genotypes ofBbss via tick transmission to American robins and white-footed mice, the Lyme disease reservoirs in North America. Among these strains, we found distinct levels of spirochete presence in the bloodstream and tissues and maintenance by these animals in a host-dependent fashion. We showed that the late stage persistence of these strains largely corresponds to bacterial survival at early infection onsets. We also demonstrated that those early survival phenotypes correspond to spirochete adhesiveness, evasion of complement-mediated killing in sera, and/or not triggering high levels of pro-inflammatory cytokines and antibodies. Our findings thus link host competence toBbss with spirochete genotypic variation of adhesiveness and inducing/escaping host immune responses, illuminating the potential mechanisms that dictate host specialization. Such information will provide a foundation for further investigation into multi-disciplinary processes driving host specialization of microparasites.AUTHOR SUMMARYHost specialization arises when microparasites adapt to a subset of available hosts, restricting the host ranges they can infect. The mechanisms and selective pressures for the evolution of host specialization remain unclear. The causative agent of Lyme disease (LD), the bacteria species complex ofBorrelia burgdorferisensu lato, is adapted to different vertebrates. However, whether such a differential host adaption also applies to each genotype within the same species is under debate. Further, the mechanisms that drive such host specialization are unclear. We thus introduced three genotypes of one LD bacteria species(B. burgdorferisensu stricto) individually via tick bite to American robins and white-footed mice, the most common LD reservoirs in North America. We found that these genotypes differed in the persistent maintenance by those reservoirs and occurred in a host-specific fashion. The ability of those bacteria for long-term maintenance was linked with their capability to attach to cells and a lack of induction of high levels of immune responses at early infection onsets. This work demonstrates the potential mechanisms that dictate host specialization of LD bacteria circulating in natural populations. Such information will pave the road to define the molecular, ecological, and evolutionary determinants that drive host-microparasite interactions.

ABSTRACT Host specialization is an ecological and evolutionary process by which a pathogen becomes differentially adapted to a subset of hosts, restricting its host range. For parasites transmitted by generalist vectors, host specialization is not expected to evolve because of the decreased survival of those parasites in inadequate hosts. Thus, parasites may develop adaptation strategies, resulting in host specialization. The causative agents of Lyme disease are multiple species of bacteria, Borrelia burgdorferi sensu lato species complex ( Bb sl), and are suitable for examining host specialization as birds and rodents were found to carry different species of these bacteria. Debate exists on whether host specialization occurs among these strains within a particular species of Bb sl, such as B. burgdorferi sensu stricto ( Bb ss). Current evidence supports some Bb ss strains are widespread in white-footed mice but others are in non-rodent vertebrates, such as birds. To recapitulate specialization in the laboratory and define the mechanisms for host specialization, we introduced different genotypes of Bb ss via tick transmission to American robins and white-footed mice, the Lyme disease reservoirs in North America. Among these strains, we found distinct levels of spirochete presence in the bloodstream and tissues and maintenance by these animals in a host-dependent fashion. We showed that the late stage persistence of these strains largely corresponds to bacterial survival at early infection onsets. We also demonstrated that those early survival phenotypes correspond to spirochete adhesiveness, evasion of complement-mediated killing in sera, and/or not triggering high levels of pro-inflammatory cytokines and antibodies. Our findings thus link host competence to Bb ss with spirochete genotypic variation of adhesiveness and inducing/escaping host immune responses, illuminating the potential mechanisms that dictate host specialization. Such information will provide a foundation for further investigation into multi-disciplinary processes driving host specialization of microparasites. AUTHOR SUMMARY Host specialization arises when microparasites adapt to a subset of available hosts, restricting the host ranges they can infect. The mechanisms and selective pressures for the evolution of host specialization remain unclear. The causative agent of Lyme disease (LD), the bacteria species complex of Borrelia burgdorferi sensu lato, is adapted to different vertebrates. However, whether such a differential host adaption also applies to each genotype within the same species is under debate. Further, the mechanisms that drive such host specialization are unclear. We thus introduced three genotypes of one LD bacteria species (B. burgdorferi sensu stricto) individually via tick bite to American robins and white-footed mice, the most common LD reservoirs in North America. We found that these genotypes differed in the persistent maintenance by those reservoirs and occurred in a host-specific fashion. The ability of those bacteria for long-term maintenance was linked with their capability to attach to cells and a lack of induction of high levels of immune responses at early infection onsets. This work demonstrates the potential mechanisms that dictate host specialization of LD bacteria circulating in natural populations. Such information will pave the road to define the molecular, ecological, and evolutionary determinants that drive host-microparasite interactions.

cellular and immunological mechanisms 2 Yi-Pin Lin*, Danielle M. Tufts, Alan P. Dupuis, II, Matthew Combs, Ashley L. 3 Marcinkiewicz, Andrew D. Hirsbrunner, Alexander J. Diaz, Jessica L. Stout, Anna M. Blom, 4 Klemen Strle, April D. Davis, Laura D. Kramer, Maria A. Diuk-Wasser* 5 Division of Infectious Diseases, Wadsworth Center, New York State Department of Health, 6 Albany, NY, USA, Department of Biomedical Sciences, State University of New York at 7 Albany, NY, USA, Department of Ecology, Evolution, and Environmental Biology, Columbia 8 University, New York, NY USA, Division of Medical Protein Chemistry, Department of 9

Abstract Urbanization exposes species to novel environments and selection pressures that may change morphological traits within a population. We investigated how the shape and size of crania and mandibles changed over time within a population of brown rats (Rattus norvegicus) living in Manhattan, New York, USA, a highly urbanized environment. We measured 3D landmarks on the cranium and mandible of 62 adult individuals sampled in the 1890s and 2010s. Static allometry explained approximately 22% of shape variation in crania and mandible datasets, while time accounted for approximately 14% of variation. We did not observe significant changes in skull size through time or between the sexes. Estimating the P‐matrix revealed that directional selection explained temporal change of the crania but not the mandible. Specifically, rats from the 2010s had longer noses and shorter upper molar tooth rows, traits identified as adaptive to colder environments and higher quality or softer diets, respectively. Our results highlight the continual evolution to selection pressures. We acknowledge that urban selection pressures impacting cranial shape likely began in Europe prior to the introduction of rats to Manhattan. Yet, our study period spanned changes in intensity of artificial lighting, human population density, and human diet, thereby altering various aspects of rat ecology and hence pressures on the skull.

Abstract As the rate of urbanization continues to increase globally, a growing body of research is emerging that investigates how urbanization shapes the movement—and consequent gene flow—of species in cities. Of particular interest are native species that persist in cities, either as small relict populations or as larger populations of synanthropic species that thrive alongside humans in new urban environments. In this study, we used genomic sequence data (SNPs) and spatially explicit individual‐based analyses to directly compare the genetic structure and patterns of gene flow in two small mammals with different dispersal abilities that occupy the same urbanized landscape to evaluate how mobility impacts genetic connectivity. We collected 215 white‐footed mice (Peromyscus leucopus) and 380 big brown bats (Eptesicus fuscus) across an urban‐to‐rural gradient within the Providence, Rhode Island (U.S.A.) metropolitan area (population =1,600,000 people). We found that mice and bats exhibit clear differences in their spatial genetic structure that are consistent with their dispersal abilities, with urbanization having a stronger effect on Peromyscus mice. There were sharp breaks in the genetic structure of mice within the Providence urban core, as well as reduced rates of migration and an increase in inbreeding with more urbanization. In contrast, bats showed very weak genetic structuring across the entire study area, suggesting a near‐panmictic gene pool likely due to the ability to disperse by flight. Genetic diversity remained stable for both species across the study region. Mice also exhibited a stronger reduction in gene flow between island and mainland populations than bats. This study represents one of the first to directly compare multiple species within the same urban‐to‐rural landscape gradient, an important gap to fill for urban ecology and evolution. Moreover, here we document the impacts of dispersal capacity on connectivity for native species that have persisted as the urban landscape matrix expands.

Effective management of rodent pests requires an ecological understanding of how they move through their environment and how those movements influence the invasion, persistence, or reinvasion of problematic colonies. Traditional methodologies used to describe rodent movement patterns, such as mark-recapture, are hindered by their time-consuming nature and limited geographic scope. As such, our understanding of how rodents interact with urban environments remains limited. Population genetic principles and tools have the capacity to greatly increase our understanding of rodent population dynamics, ecological relationships, and movements across space, but this field is often unapproachable to non-scientist pest management professionals (PMPs). In this commentary, we aim to promote collaborative and integrative rodent pest management by introducing relevant population genetic principles, providing examples of their applications in studies of urban brown rats (Rattus norvegicus), and proposing future initiatives that link scientific, private, and government entities. We reinterpret results from a 2018 study of brown rats in Vancouver, British Columbia, Canada to show how genetic relationships among individual brown rats can be used to understand the geographic distribution of genetic clusters (i.e., colonies), natural barriers to migration, and the spatial scale of dispersal. While the 2018 study originally aimed to describe patterns of population genetic structure to understand the influence of urban landscapes on rats, here we describe how these results can be exploited by PMPs to directly inform the creation of management units and decrease the likelihood of rapid post-treatment reinvasion. Further, we discuss the difficulties inherent in population genetic studies and the potential for high-quality model sites to develop generalizable strategies. Overall, we hope to expand the toolbox of PMPs, foster collaboration, and move toward more informed and sustainable management strategies.

Urbanization often substantially influences animal movement and gene flow. However, few studies to date have examined gene flow of the same species across multiple cities. In this study, we examine brown rats (Rattus norvegicus) to test hypotheses about the repeatability of neutral evolution across four cities: Salvador, Brazil; New Orleans, USA; Vancouver, Canada; and New York City, USA. At least 150 rats were sampled from each city and genotyped for a minimum of 15 000 genome-wide single nucleotide polymorphisms. Levels of genome-wide diversity were similar across cities, but varied across neighbourhoods within cities. All four populations exhibited high spatial autocorrelation at the shortest distance classes (less than 500 m) owing to limited dispersal. Coancestry and evolutionary clustering analyses identified genetic discontinuities within each city that coincided with a resource desert in New York City, major waterways in New Orleans, and roads in Salvador and Vancouver. Such replicated studies are crucial to assessing the generality of predictions from urban evolution, and have practical applications for pest management and public health. Future studies should include a range of global cities in different biomes, incorporate multiple species, and examine the impact of specific characteristics of the built environment and human socioeconomics on gene flow.

Range expansion has genetic consequences expected to result in differentiated wave-front populations with low genetic variation and potentially introgression from a local species. The northern expansion of Peromyscus leucopus in southern Quebec provides an opportunity to test these predictions using population genomic tools. Our results show evidence of recent and post-glacial expansion. Genome-wide variation in P. leucopus indicates two post-glacial lineages are separated by the St. Lawrence River, with a more recent divergence of populations isolated by the Richelieu River. In two of three transects we documented northern populations with low diversity in at least one genetic measure, although most relationships were not significant. Consistent with bottlenecks and allele surfing during northward expansion, we document a northern-most population with low nucleotide diversity, divergent allele frequencies and the most private alleles, and observed heterozygosity indicates outcrossing. Ancestry proportions revealed putative hybrids of P. leucopus and P. maniculatus. A formal test for gene flow confirmed secondary contact, showing that a reticulate population phylogeny between P. maniculatus and P. leucopus was a better fit to the data than a bifurcating model without gene flow. Thus, we provide the first genomic evidence of gene flow between this pair of species in natural populations. Understanding the evolutionary consequences of secondary contact is an important conservation concern as climate-induced range expansions are expected to result in new hybrid zones between closely related species.

Human commensal species such as rodent pests are often widely distributed across cities and threaten both infrastructure and public health. Spatially explicit population genomic methods provide insights into movements for cryptic pests that drive evolutionary connectivity across multiple spatial scales. We examined spatial patterns of neutral genomewide variation in brown rats (Rattus norvegicus) across Manhattan, New York City (NYC), using 262 samples and 61,401 SNPs to understand (i) relatedness among nearby individuals and the extent of spatial genetic structure in a discrete urban landscape; (ii) the geographic origin of NYC rats, using a large, previously published data set of global rat genotypes; and (iii) heterogeneity in gene flow across the city, particularly deviations from isolation by distance. We found that rats separated by ≤200 m exhibit strong spatial autocorrelation (r = .3, p = .001) and the effects of localized genetic drift extend to a range of 1,400 m. Across Manhattan, rats exhibited a homogeneous population origin from rats that likely invaded from Great Britain. While traditional approaches identified a single evolutionary cluster with clinal structure across Manhattan, recently developed methods (e.g., fineSTRUCTURE, sPCA, EEMS) provided evidence of reduced dispersal across the island's less residential Midtown region resulting in fine‐scale genetic structuring (FST = 0.01) and two evolutionary clusters (Uptown and Downtown Manhattan). Thus, while some urban populations of human commensals may appear to be continuously distributed, landscape heterogeneity within cities can drive differences in habitat quality and dispersal, with implications for the spatial distribution of genomic variation, population management and the study of widely distributed pests.