Aims: To characterize the genetic relatedness, phenotypic and genotypic antimicrobial resistance and plasmid content of 80 Salmonella Infantis strains isolated from food, humans and veterinary sources from 2013 to 2018 in Brazil. Methods and results: Pulsed-field gel electrophoresis and single-nucleotide polymorphism analysis showed major clusters containing 50% and 38.8% of the strains studied respectively. Multilocus sequence typing assigned all strains to ST32. Disk-diffusion revealed that 90% of the strains presented resistant or intermediate resistant profiles and 38.8% displayed multidrug resistance. Resistance genes for aminoglycosides (aac(6′)-Iaa; aadA12; aph(3″-Ib; aph(6)-Id), β-lactams (blaTEM-1; blaCTX-M- 8; blaCMY-2), trimethoprim (dfrA8), tetracycline (tet(A)), amphenicols (floR), sulfonamide (sul2), efflux pumps (mdsA; mdsB), chromosomal point mutations in gyrB, parC, acrB and pmrA were detected. Strains harboured IncI, IncF, IncX, IncQ, IncN and IncR plasmids. Conclusions: The presence of a prevalent S. Infantis subtype in Brazil and the high antimicrobial resistance rates reinforced the potential hazard of this serovar for the public health and food safety fields. Significance and Impact of the Study: This is the first study characterizing a large set of S. Infantis from Brazil by whole-genome sequencing, which provided a better local and global comprehension about the distribution and characteristics of this serovar of importance in the food, human and veterinary fields.