Understanding how vaccine hesitancy relates to online behavior is crucial for addressing current and future disease outbreaks. We combined survey data measuring attitudes toward the COVID-19 vaccine with Twitter data in two studies (N1 = 464 Twitter users, N2 = 1,600 Twitter users) with preregistered hypotheses to examine how real-world social media behavior is associated with vaccine hesitancy in the United States (US) and the United Kingdom (UK). In Study 1, we found that following the accounts of US Republican politicians or hyper-partisan/low-quality news sites were associated with lower confidence in the COVID-19 vaccine—even when controlling for key demographics such as self-reported political ideology and education. US right-wing influencers (e.g. Candace Owens, Tucker Carlson) had followers with the lowest confidence in the vaccine. Network analysis revealed that participants who were low and high in vaccine confidence separated into two distinct communities (or “echo chambers”), and centrality in the more right-wing community was associated with vaccine hesitancy in the US, but not in the UK. In Study 2, we found that one’s likelihood of not getting the vaccine was associated with retweeting and favoriting low-quality news websites on Twitter. Altogether, we show that vaccine hesitancy is associated with following, sharing, and interacting with low-quality information online, as well as centrality within a conservative-leaning online community in the US. These results illustrate the potential challenges of encouraging vaccine uptake in a polarized social media environment.