Abstract Background Pancreatic cancer (PC) presents a significant challenge in oncology because of its late-stage diagnosis and limited treatment options. The inadequacy of current screening methods has prompted investigations into stool-based assays and microbial classifiers as potential early detection markers. The gut microbiota composition of PC patients may be influenced by population differences, thereby impacting the accuracy of disease prediction. However, comprehensive profiling of the PC gut microbiota and analysis of these cofactors remain limited. Therefore, we analyzed the stool microbiota of 33 Finnish and 50 Iranian PC patients along with 35 Finnish and 34 Iranian healthy controls using 16S rRNA gene sequencing. We assessed similarities and differences of PC gut microbiota in both populations while considering sociocultural impacts and generated a statistical model for disease prediction based on microbial classifiers. Our aim was to expand the current understanding of the PC gut microbiota, discuss the impact of population differences, and contribute to the development of early PC diagnosis through microbial biomarkers. Results Compared with healthy controls, PC patients presented reduced microbial diversity, with discernible microbial profiles influenced by factors such as ethnicity, demographics, and lifestyle. PC was marked by significantly higher abundances of facultative pathogens including Enterobacteriaceae, Enterococcaceae, and Fusobacteriaceae, and significantly lower abundances of beneficial bacteria. In particular, bacteria belonging to the Clostridia class, such as butyrate-producing Lachnospiraceae, Butyricicoccaceae, and Ruminococcaceae, were depleted. A microbial classifier for the prediction of pancreatic ductal adenocarcinoma (PDAC) was developed in the Iranian cohort and evaluated in the Finnish cohort, where it yielded a respectable AUC of 0.88 (95% CI 0.78, 0.97). Conclusions This study highlights the potential of gut microbes as biomarkers for noninvasive PC screening and the development of targeted therapies, emphasizing the need for further research to validate these findings in diverse populations. A comprehensive understanding of the role of the gut microbiome in PC could significantly enhance early detection efforts and improve patient outcomes.