We retrospectively collected PD patients with a performance of bile culture between 2007 and 2019 in our institute. As to bile culture, we used a swab to do intraoperative bile cultures after transection of the CBD. IAA was defined as the documental bacteriological culture from either a turbid discharge from the intraoperatively placed drain in patients with a clinical picture consistent with infection or a postoperative fluid collection managed by CT-guided placement of drains. A total of 1244 PD patients were identified, and 539 (43.3%) subjects with bile sampling were included for analysis. Among these study patients, 433 (80.3%) developed bile contamination (positive bile culture). Bile contamination showed a significantly higher rate of IAA compared to non-bile contamination (17.1% vs. 0.9%, p < 0.001). The rate of co-shared microorganisms in both bile and abscess was 64.1%. On the multivariate analysis, age and specific bile microorganisms (Enterococcus species, Escherichia Coli, Streptococcus species, Citrobacter species, and Candida) are significantly associated with development of IAA. Specific bile microorganisms are the highly significant factors associated with development of IAA. The strategy to prevent bile spillage during PD should be considered to minimize afterward contamination of the abdominal cavity and prevent IAA.