Background: Campylobacter spp. have been reported as a common cause of gastroenteritis in humans in many countries. However, in Brazil there is insufficient data to estimate the impact of Campylobacter in public health. In light of the importance of this foodborne pathogen, the aim of this study was to perform comparative analyses on 80 Brazilian Campylobacter coli genomes isolated from human feces, animals, the environment, and food. Methods include Average Nucleotide Identity (ANI), Gegenees, genomic plasticity, presence of pathogenicity, resistance, and metabolic islands. In addition, virulence analysis in Galleria mellonella were also performed for 18 selected C. coli strains. Results: The ANI values confirmed that all strains belonged to the C. coli species. Phylogenetic analyses demonstrated the evolutionary relationships among the studied strains, highlighting the genetic diversity among them. The differences in shared and deleted regions of the studied genomes were demonstrated, with 16 genomic islands identified, including 4 metabolic islands, 4 resistance islands, and 8 pathogenicity islands. We detected genes associated with chemotaxis, exotoxin production, antimicrobial resistance, stress response, defense mechanisms, and intracellular survival among these islands, highlighting the pathogenic potential of these strains. Two strains isolated from human and one from animal showed high virulence, killing 100% of Galleria mellonella larvae. Two strains isolated from the environment and two isolated from food killed 70–90% of the larvae and were classified as virulent. Three strains isolated from animal, two from human, two from the environment and one from food killed 30% to 60% of the larvae and were considered of intermediate virulence. Campylobacter jejuni ATCC 33291, one strain isolated from human and one from food killed 10 to 20% of the larvae and were considered of low virulence. One strain isolated from food did not kill any larvae and was considered avirulent. Conclusions: The results obtained highlighted the genetic diversity, pathogenic and virulence potential of many of the C. coli strains studied, contributing for a more complete characterization of this important pathogen recognized as a cause of human gastroenteritis.