The Po river plain (Northern Italy) hosts artificial, lowland springs locally known as fontanili, which provide important ecosystem services in an area dominated by intensive agricultural activities. Here we present a study carried out in 50 springs. Each spring was visited once from October 2015 to January 2016. The sampled sites were selected to include springs studied in 2001 and 2004, to evaluate changes in water quality and ostracod assemblages that possibly occurred over a period of 10–15 years, and explore the relationships between ostracod community composition and water physical and chemical variables. Our results showed a decrease in the chemical water quality especially, in springs south of the Po river, evidenced by high nitrate levels. Most of the studied springs showed a relevant decrease in dissolved reactive silica, probably related to recent transformations of either agricultural practices or crop typology. Ostracods were mostly represented by common and tolerant species, and communities were characterized by low alpha diversity and high species turnover. Water temperature and mineralization level were the most influential variables in structuring the ostracod communities. We stress the need to implement conservation and restoration measures for these threatened ecosystems, to regain their role as ecosystem services providers.