Background: Group B Streptococcus (GBS) is a predominant cause of early-onset neonatal sepsis globally; however, the impact of this organism on the health of newborns in South Asia is debated, due in part to a paucity of community-based assessments. We estimated the prevalence and serotypes of GBS colonization of the umbilical cord stump and the association of colonization with mortality in a population-based cohort of newborns in rural Sylhet District, Bangladesh. Methods: Umbilical cord swabs were collected from 646 newborns up to 3 times within the first week after birth (ages <24 hours, ∼3 days, ∼6 days) and processed to identify GBS serotypes. Results: GBS was isolated from 6.3% (41/646) of newborns. Sixty-one percent of the GBS colonization occurred in neonates within 24 hours of delivery. Serotypes VII (37.1%, n = 13/36) and Ia (33.3%, n = 12/36) were the most predominant colonizing GBS isolates. Other detected serotypes were Ib (11.1%, n = 4/36), II (11.1%, n = 4/36), V (5.6%, n = 2/36) and VI (2.8%, n = 1/36). Mortality risk among newborns with GBS colonization was 6.6 (95% confidence interval: 2.1-20.4) times higher than for those without GBS. Conclusions: The overall prevalence of GBS colonization was lower than in settings, where GBS is a predominant etiology of neonatal illness. In addition, the GBS serotype distribution differed from that reported in the developed part of the world. However, further studies are needed to understand the true burden of GBS-related illness. Mortality risk was substantially increased in the presence of GBS on the umbilical stump, providing support for chlorhexidine antisepsis to the cord to prevent colonization of invasive pathogens.