Geographically expanding and invading ticks are a global concern. The Asian longhorned tick (ALT, Haemaphysalis longicornis) was introduced to the mid-Atlantic US between 2010 and 2017 and recently invaded Ohio, an inland state. To date, ALTs in the US have been associated with livestock exsanguination and transmission of the agent of bovine theileriosis. To inform management, studies describing tick ecology and epidemiology of associated disease agents are critical. In this study, we described phenology, habitat and host associations, and tested for agents of medical and veterinary concern at the site of the first known established ALT population in Ohio, where pesticide treatment was applied in early fall 2021. In spring-fall 2022, we sampled wildlife (small mammals) and collected ticks from forest, edge, and grassland habitats. We also opportunistically sampled harvested white-tailed deer at nearby processing stations and fresh wildlife carcasses found near roads. Field-collected ALTs were tested for five agents using real-time PCR. We found that ALT nymphs emerged in June, followed by adults, and concluded with larvae in the fall. ALTs were detected in all habitats but not in wildlife. We also found a 4.88% (2/41) prevalence of Anaplasma phagocytophilum across ALT adults and nymphs. Host and habitat associations were similar to other studies in the eastern United States, but two potential differences in phenology were identified. Whether ALTs will acquire more endemic disease agents requires further investigations. Our findings provide the first evidence regarding ALT life history from the Midwest region of the United States and can inform exposure risk and guide integrated management.