This article seeks to contribute to the study of migration and religion in two EU countries, Croatia and Italy, by examining the impact of religiosity and cultural identification on negative attitudes toward immigrants. In many European societies, the increasing diversification within different levels of society stemming from recent migrations has turned immigrants’ reception and integration into a key issue, whereby migrants are often perceived as a threat to the dominant religion and culture, thus aggravating the process of migrant integration within society. Our article follows recent empirical research on migration and religion, which determined that higher levels of religiosity are positively correlated with negative out-group attitudes. Conducting quantitative research in Croatia (N = 603) and Italy (N = 714) and based on the analysis of primary data, firstly, we assess whether there is an association between negative attitudes towards immigrants depending on different degrees of religiosity and levels of cultural identification. Secondly, we examine the differences of the socio-religious contexts of Croatia and Italy, with a focus on the interplay between religion, national identity, and migration patterns. In line with this, our research shows that religiosity has the largest influence on negative attitudes toward immigrants, implying that higher levels of religiosity result in higher levels of negative attitudes toward immigrants. Furthermore, the results of our research show that Croatian participants have more negative attitudes toward immigrants than Italian participants, whereby Roman Catholic participants in both countries are more negative than non-religiously declared participants.