Serpa cheese is one of the traditional regional Portuguese cheeses having the Protected Denomination of Origin (PDO) designation. This study investigated the bacterial community in the traditional Portuguese Serpa cheese. The microorganisms identified at the end of ripening (30 days) mainly were lactic acid bacteria (LAB). Lactobacillus paracasei/Lactobacillus casei was the main species in cheese from PDO registered industries, whereas in non‐PDO registered industries Lactobacillus brevis was highlighted, among other LAB. Enterobacteriaceae species were detected at 20% to 40% of the total isolates. The results obtained by high‐throughput sequencing analysis confirmed that LAB was the main microbial group, with Lactococcus genus contributing to approximately 40% to 60% of the population, followed by Leuconostoc and Lactobacillus. The Enterobacteriaceae family was also important. The differences between bacterial communities from PDO and non‐PDO registered industries suggest that the lack of regulation of the cheese‐making practices may influence unfavorably. The new knowledge about bacterial diversity in Serpa cheese could be useful to set up new ripening conditions, which favor the development of desirable microorganisms. Practical Application The control of the manufacturing process of traditional cheeses can be improved through the knowledge of the bacterial diversity that develops. Thus, the growth of desirable microorganisms can be promoted to homogenize the final product.