(1) Background: The global health crisis caused by the coronavirus disease (COVID-19) pandemic has led to extreme overloading of different public healthcare systems worldwide. The Spanish Public Healthcare System is one of them. This study aimed to conduct a comparative cost analysis to assess the impact of the COVID-19 pandemic on small- and medium-sized regional hospitals in Andalusia (Spain). (2) Methods: This comparative, multicentre, observational, and retrospective study was designed to perform a comparative cost analysis between the Alto Guadalquivir Health Agency (AGHA) and Poniente University Hospital (PUH), both of which belong to the Spanish Public Health System (PHS). The data included in this study corresponds to the total costs by area and year incurred by the 61,335 patients from both healthcare institutions (AGHA = 36,110; PUH = 25,225) in the areas of hospital emergency service (HES), hospitalisation, and intensive care unit (ICU), during the 24 months of the study period (from 1 January 2019 to 31 December 2020). (3) Results: The analysis results showed a significant increase in costs incurred in 2020 for HES relative to those incurred in 2019 for both AGHA (+14%; p < 0.003) and PUH (+36%; p = 0.002). Additionally, costs incurred for ICU increased significantly in 2020 relative to those incurred in 2019 for both AGHA (+30%; p = 0.003) and PUH (+46%; p = 0.002). Hospitalisation costs for AGHA also increased significantly (+9%; p < 0.012) in 2020 versus those obtained in 2019; however, no significant differences were found for PUH (+6%; p = 1) in the same period analysed. However, the number of patients treated in the areas of HES, hospitalisation, and ICU was significantly reduced throughout 2020 compared to 2019. (4) Conclusions: Our findings show that the costs incurred during 2020 in the regional hospitals of Andalusia (Spain) increased significantly in most of the parameters analysed relative to those incurred in the year before the pandemic (i.e., 2019).