Enterococcus faeciumis sometimes used in food production; however, its acquisition of antibiotic resistance has become an alarming health concern. TheE. lactisspecies is closely related toE. faeciumand has good probiotic potential. This study aimed to investigate the antibiotic resistance ofE. lactis.We analyzed the antibiotic resistance phenotype and whole-genome sequences of 60E. lactisisolates (23, 29, and 8 isolates from dairy products, Rice wine Koji, and human feces, respectively). These isolates showed varying degree of resistance to 13 antibiotics, and were sensitive to ampicillin and linezolid. TheE. lactisgenomes carried only a subset of commonly reported antibiotic resistance genes (ARGs) inE. faecium. Five ARGs were detected across the investigatedE. lactis, including two universally present genes (msrCandAAC(6′)-Ii) and three rarely detected ARGs (tet(L),tetM, andefmA). To identify other undescribed antibiotic resistance-encoding genes, a genome-wide association study was performed, returning 160 potential resistance genes that were associated with six antibiotics, namely chloramphenicol, vancomycin, clindamycin, erythromycin, quinupristin-dalfopristin, and rifampicin. Only around one-third of these genes encode known biological functions, including cellular metabolism, membrane transport, and DNA synthesis. This work identified interesting targets for future study of antibiotic resistance inE. lactis. The fact that the lower number of ARGs present inE. lactissupports that it may be an alternative toE. faecalisfor use in the food industry. Data generated in this work is of interest to the dairy industry.