Genome analysis of Streptococcus spp. isolates from Animals in Pre-Antibiotic Era with Respect to Antibiotic Susceptibility and Virulence Gene Profiles

Abstract Lyophilized Streptococcus spp. isolates (n = 50) from animal samples submitted to the diagnostic laboratory at the University of Connecticut in the 1940s were revivified to investigate the genetic characteristics using whole-genome sequencing (WGS). The Streptococcus spp. isolates were identified as follows; S. agalactiae (n = 14), S. dysgalactiae subsp. dysgalactiae (n = 10), S. dysgalactiae subsp. equisimils (n = 5), S. uberis (n = 8), S. pyogenes (n = 7), S. equi subsp. zooepidemicus (n = 4), S. oralis (n = 1), and S. pseudoporcinus (n = 1). We identified sequence types (ST) of S. agalactiae, S. dysgalactiae, S. uberis, S. pyogenes, and S. equi subsp. zooepidemicus and reported ten novel sequence types of those species. Phylogenetic analysis revealed high genetic diversity of Streptococcus spp. in animals in the United States in the 1940s. WGS analysis revealed that none of Streptococcus spp. carried acquired antibiotic resistance genes but all S. agalactiae isolates carried the macrolide resistance gene (mre(A)). However, tetracycline resistance was observed in four out of 14 S. dysgalactiae isolates and in one S. equi subsp. zooepidemicus isolate. This data highlights that antimicrobial resistance is pre-existed in nature before the use of antibiotics. Virulence gene profile analysis revealed that the hasC gene was observed in all Streptococcus spp. isolates of this study, and none of the Streptococcus spp. isolates in this study harbored the cba and clpP genes. This study provides an invaluable resource for studying the evolutionary aspects of antibiotic resistance acquisition and virulence in Streptococcus spp.