Bacteria develop resistance to antibiotics naturally, but the inappropriate and widespread use of antibiotics in humans and animals has made antimicrobial resistance one of the biggest threats to modern medicine. Raw milk cheese can represent an important source of antimicrobial resistance. Thus, the objective of this study was to evaluate the prevalence and sensitivity of Escherichia coli isolated from artisanal cheese made from raw milk produced in Minas Gerais, Brazil. E. coli counts were determined using the most probable number method. An antibiogram was performed using the disk diffusion method, following the protocol described by the Brazilian Committee on Antimicrobial Susceptibility Testing (BrCAST) for 14 antibiotics of nine classes. E. coli was detected in 35 (71.4%) of the samples, with populations between 0.56 to 4.87 log (NMP/g) of cheese. The presence of E. coli resistant to multiple antimicrobials was more frequent in cheeses, with an E. coli population below the levels established by regulatory limits. Only four samples (11.4%) had all E. coli isolates susceptible to the 14 antimicrobials evaluated. The results showed the heterogeneity of antimicrobial resistance in E. coli between the producing regions of Minas artisanal cheese. Multidrug resistance was detected in 29% of the E. coli isolates and in almost 40% (38.8%) of the cheese samples. The frequency of multidrug-resistant (MDR) isolates was different between the production regions (p < 0.05). The presence of MDR E. coli in cheese from region D was 14, 4, and 20 times more likely than in cheese from regions A, B, and C, respectively. A multiple antibiotic resistance index of 0.200 predicted the presence of MDR E. coli in raw milk artisanal cheese with 99% probability. In conclusion, artisanal cheese can act as sources of MDR E. coli to colonize the human gastrointestinal tract.