Objectives: Data on the prevalence of chronic diseases and their relationship with sickness absence in the Italian public employees are rather scarce. Therefore, in the first place, we assessed the distribution of chronic diseases in the employees of the University of Ferrara. As a next step, we investigated the possible associations between each chronic disease and cumulative days of all-cause sickness absence, and finally we investigated the odds ratio of each single chronic disease on sickness absence. Material and Methods: A total of 514 employees, 269 sick-listed and 245 not sick-listed in 2012, were studied. Demographical/clinical characteristics and chronic diseases were obtained from all study participants during medical surveillance procedures. Sickness absence days and job seniority data were obtained from the administrative office. Results: Gastrointestinal and psychiatric diseases were the most reported in the sick-listed sample (p = 0.01 and p = 0.02, respectively, compared to the not sick-listed). In the interquantile regression analysis, the sickness absence days were associated with psychiatric diseases (β = 65.1, 95% CI: 13.2-117.1, p = 0.01) and with the presence of 2 or more chronic diseases (β = 23.3, 95% CI: 4.5-42, p = 0.02). Furthermore, the logistic regression analysis showed that the odds of sickness absence were increased 2 fold by psychiatric diseases (OR = 2.2, 95% CI: 1.01-4.93, p = 0.04), and gastrointestinal diseases (OR = 1.9, 95% CI: 1.07-3.42, p = 0.02) and, to a lesser extent, by high body mass index (OR = 1.05, 95% CI: 1-1.11, p = 0.03). Conversely, female gender reduced by half the odds of sickness absence (OR = 0.5, 95% CI: 0.3-0.8, p = 0.04). Conclusions: This study highlights the relevant association between chronic diseases and sickness absence in Italian public employees. Our findings indicate the importance of considering the health status when designing preventive interventions aimed at decreasing sickness absences in this population.