Environmental Reservoirs of Microbial Contamination in University Food Services: A Large-Scale Study in Northern Portugal

(1) Background: University food service establishments are complex environments, where high turnover and handling practices create conditions for microbial persistence. Food-contact surfaces (FCSs) and handlers’ hands (FHs) function as dynamic reservoirs, facilitating the circulation of contaminants within these institutional settings. This study aimed to characterise the microbiological contamination of FCSs and FHs in university food service establishments in Northern Portugal and to evaluate their role as interconnected environmental reservoirs within the indoor built environment. (2) Methods: A total of 590 samples were analysed from two universities in Northern Portugal (L1, L2), comprising 380 FCS and 210 FH samples. Aerobic colony counts (ACCs), Enterobacteriaceae, and Moulds and yeasts (MYs) were analysed according to ISO methods. FH samples were additionally screened for Escherichia coli and Staphylococcus spp. (3) Results: Overall, 35.5% of FCSs were classified as non-compliant, according to microbial criteria based on guideline values from the National Health Institute Dr. Ricardo-Jorge (INSA), with non-compliance primarily driven by elevated ACCs and MYs. Based on a Generalised Linear Model (GLM), establishment types (canteens vs. cafes) were associated with Enterobacteriaceae levels (p = 0.016), whereas ACCs and MYs were not significantly associated with district, establishment type, or functional surface category (p > 0.05). Differences between left and right hands showed small effect sizes, and location was a highly significant determinant of hand hygiene acceptability. (4) Conclusions: FCSs and FHs act as relevant contamination reservoirs in these settings. The results indicate that Enterobacteriaceae levels on FCSs differed between establishment types, while ACCs and MYs showed no significant variation across the evaluated environmental factors. Marked differences in hand hygiene acceptability between campuses support the implementation of targeted interventions, including the optimisation of cleaning and disinfection protocols, the structured training of food handlers, and the routine microbiological monitoring of surfaces and hands to improve institutional food safety.