Abstract: BACKGROUND The extent of urban areas is increasing around the world, and most humans now live in cities. Urbanization results in dramatic environmental change, including increased temperatures, more impervious surface cover, altered hydrology, and elevated pollution. Urban areas also host more non-native species and reduced abundance and diversity of many native species. These environmental changes brought by global urbanization are creating novel ecosystems with unknown consequences for the evolution of life. Here, we consider how early human settlements led to the evolution of human commensals, including some of the most notorious pests and disease vectors. We also comprehensively review how contemporary urbanization affects the evolution of species that coinhabit cities. ADVANCES A recent surge of research shows that urbanization affects both nonadaptive and adaptive evolution. Some of the clearest results of urban evolution show that cities elevate the strength of random genetic drift (stochastic changes in allele frequencies) and restrict gene flow (the movement of alleles between populations due to dispersal and mating). Populations of native species in cities often represent either relicts that predate urbanization or populations that established after a city formed. Both scenarios frequently result in a loss of genetic diversity within populations and increased differentiation between populations. Fragmentation and urban infrastructure also create barriers to dispersal, and consequently, gene flow is often reduced among city populations, which further contributes to genetic differentiation between populations. The influence of urbanization on mutation and adaptive evolution are less clear. A small number of studies suggest that industrial pollution can elevate mutation rates, but the pervasiveness of this effect is unknown. A better studied phenomenon are the effects of urbanization on evolution by natural selection. A growing number of studies show that plant and animal populations experience divergent selection between urban and nonurban environments. This divergent selection has led to adaptive evolution in life history, morphology, physiology, behavior, and reproductive traits. These adaptations typically evolve in response to pesticide use, pollution, local climate, or the physical structure of cities. Despite these important results, the genetic basis of adaptive evolution is known from only a few cases. Most studies also examine only a few populations in one city, and experimental validation is rare. OUTLOOK The study of evolution in urban areas provides insights into both fundamental and applied problems in biology. The thousands of cities throughout the world share some features while differing in other aspects related to their age, historical context, governmental policies, and local climate. Thus, the phenomenon of global urbanization represents an unintended but highly replicated global study of experimental evolution. We can harness this global urban experiment to understand the repeatability and pace of evolution in response to human activity. Among the most important unresolved questions is, how often do native and exotic species adapt to the particular environmental challenges found in cities? Such adaptations could be the difference as to whether a species persists or vanishes from urban areas. In this way, the study of urban evolution can help us understand how evolution in populations may contribute to conservation of rare species, and how populations can be managed to facilitate the establishment of resilient and sustainable urban ecosystems. In a similar way, understanding evolution in urban areas can lead to improved human health. For example, human pests frequently adapt to pesticides and evade control efforts because of our limited understanding of the size of populations and movement of individuals. Applied evolutionary studies could lead to more effective mitigation of pests and disease agents. The study of urban evolution has rapidly become an important frontier in biology, with implications for healthy and sustainable human populations in urban ecosystems.