Purpose of investigation: This study aimed to ascertain the colonization rate of Group B Streptococcus (GBS) in the birth canals of late-pregnant women, and to demonstrate the characteristics of the origins of antibiotic-resistance. Materials and Methods: A prospective survey of pregnant women from 2017 to 2019 at a university-affiliated hospital in China was conducted. A total of 275 pregnant women with a pregnancy between 35 and 37 weeks were included in the study. Vaginal secretions and rectal swabs were tested by GBS separation to identify cultures, and an agar doubling dilution method was performed to assess drug sensitivity. Results: In 275 pregnant women, 20 cases were GBS screen-positive for the vaginal swabs with a GBS colonization rate of 7.27%. In addition, 16 isolates from the rectal swabs were also GBS screen-positive. Of 20 isolates from the vaginal swabs, 90.0% were resistant to tetracycline, 70.0% were resistant to erythromycin, 40.0% were resistant to clindamycin, 40.0% were resistant to levofloxacin and 30.0% resistant to chloromycetin. All isolates were susceptible to penicillin G, vancomycin, and linezolid. Conclusions: The higher prevalence of GBS colonization among pregnant women indicated the critical need for screening GBS colonization in late-pregnant women in the region. The study revealed the resistant patterns of GBS colonization and inferred a possible mechanism of the GBS resistance to antimicrobial agents used as animal growth promotion substances.