Urbanization can influence animal traits, including immunity and gut microbiota. This idea has been studied in well-established urban areas, but few studies have determined the effect of early-stage urbanization on the gut microbiota and its relationship with immunity. Over the past several decades, the Galapagos Islands have seen rapid resident human population growth and tourist activity, leading to varying levels of human activity of four of thirteen Islands. Consequently, diet, gut microbiota, and immunity of endemic animals, such as Darwin's finches, may have changed. The goal of this study was to determine the effect of land use on the immune response, gut microbiota, and body measurements of Darwin's finches early-on in the rapid increase in human activity in the Islands. Specifically, we compared proxies of the immune response (haptoglobin levels, lysozyme activities, complement antibody levels, and natural antibody levels), gut microbiota (bacterial diversity, community structure and membership, and relative abundance of bacterial taxa), and body measurements (body mass, tarsus length, and scaled mass index) across undeveloped, agricultural, and urban areas in 2008 for medium ground finches (Geospiza fortis) and small ground finches (G. fuliginosa). We found that lysozyme activity was lower and bacterial diversity was higher in urban areas compared to non-urban areas across both finch species. For small ground finches, relative abundances of bacterial genera (Pseudoxanthomonas, Cloacibacterium, and Dietzia spp.) were higher in urban areas compared to non-urban areas, but this pattern was not observed in medium ground finches. Medium ground finches were smaller in undeveloped areas compared to the other two areas, but body measurements of small ground finches did not differ across areas. Our historical data provides an interesting comparison to more recent data and demonstrates that even early human activity can influence traits of organisms.