Hyaluronidase Impairs Neutrophil Function and Promotes Group BStreptococcusInvasion and Preterm Labor in Nonhuman Primates

Abstract

Invasive bacterial infections during pregnancy are a major risk factor for preterm birth, stillbirth, and fetal injury. Group B streptococci (GBS) are Gram-positive bacteria that asymptomatically colonize the lower genital tract but infect the amniotic fluid and induce preterm birth or stillbirth. Experimental models that closely emulate human pregnancy are pivotal for the development of successful strategies to prevent these adverse pregnancy outcomes. Using a unique nonhuman primate model that mimics human pregnancy and informs temporal events surrounding amniotic cavity invasion and preterm labor, we show that the animals inoculated with hyaluronidase (HylB)-expressing GBS consistently exhibited microbial invasion into the amniotic cavity, fetal bacteremia, and preterm labor. Although delayed cytokine responses were observed at the maternal-fetal interface, increased prostaglandin and matrix metalloproteinase levels in these animals likely mediated preterm labor. HylB-proficient GBS dampened reactive oxygen species production and exhibited increased resistance to neutrophils compared to an isogenic mutant. Together, these findings demonstrate how a bacterial enzyme promotes GBS amniotic cavity invasion and preterm labor in a model that closely resembles human pregnancy. IMPORTANCE Group B streptococci (GBS) are bacteria that commonly reside in the female lower genital tract as asymptomatic members of the microbiota. However, during pregnancy, GBS can infect tissues at the maternal-fetal interface, leading to preterm birth, stillbirth, or fetal injury. Understanding how GBS evade host defenses during pregnancy is key to developing improved preventive therapies for these adverse outcomes. In this study, we used a unique nonhuman primate model to show that an enzyme secreted by GBS, hyaluronidase (HylB) promotes bacterial invasion into the amniotic cavity and fetus. Although delayed immune responses were seen at the maternal-fetal interface, animals infected with hyaluronidase-expressing GBS exhibited premature cervical ripening and preterm labor. These observations reveal that HylB is a crucial GBS virulence factor that promotes bacterial invasion and preterm labor in a pregnancy model that closely emulates human pregnancy. Therefore, hyaluronidase inhibitors may be useful in therapeutic strategies against ascending GBS infection.

COVID-19 generates hyaluronan fragments that directly induce endothelial barrier dysfunction

“…In bacterial sepsis, glycocalyx degradation associates with the presence and severity of disease and is a causative element in recruitment of immune cells, barrier disruption, and development of ARDS (31)(32)(33)(34). The glycocalyx is degraded at the cell surface by the activity of endo-β-glucuronidases which regulate the availability of HA or HS chains for receptor binding by cleaving these polymers and thereby control several important ligand-binding interactions with known roles in leukocyte recruitment, neutrophil function , endothelial behavior, and cytokine and growth factor release (33,(35)(36)(37)(38)(39). Hyaluronidases predominantly degrade HA while heparanase predominantly degrades HS, and both exhibit limited activity toward CS which has no known specific degradation enzyme in mammals (40,41).…”

Bacterial and Host Determinants of Group B Streptococcal Vaginal Colonization and Ascending Infection in Pregnancy

“…However, GBS expressing hyaluronidase (HylB) inhibit this mechanism by interfering with toll-like receptor 2 and 4 signaling. Downstream effects of this immune dampening suppress ROS production and facilitate GBS survival (Coleman et al, 2021).…”

Non-human Primate Models to Investigate Mechanisms of Infection-Associated Fetal and Pediatric Injury, Teratogenesis and Stillbirth

“…To overcome these challenges, a chronically catheterized pregnant NHP model was developed to monitor GBS-induced disease progression, immune response, and pregnancy outcomes following an experimental GBS inoculation ( Gravett et al, 1994 ; Adams Waldorf et al, 2011a ; Boldenow et al, 2016 ; Coleman et al, 2020 , 2021 ). This model is highly translational based on several characteristics: (1) inoculation of GBS in the choriodecidual space where GBS is hypothesized to first invade the placenta, (2) serial sampling of maternal and fetal blood and amniotic fluid, (3) monitoring of uterine contractions and fetal heart rate over time, and (4) administration of antibiotics or immunotherapeutics to prevent preterm birth or fetal injury ( Figure 5 ).…”

“…GBS HylB breaks down hyaluronan (HA) to into HA disaccharides that block recognition and subsequent signaling by toll-like receptor (TLR) 2 and TLR4 ( Kolar et al, 2015 ). NHP infected with GBS that highly express HylB (strain GB37; Gendrin et al, 2018 ) consistently exhibited microbial invasion of the amniotic cavity, fetal sepsis and preterm labor ( Kolar et al, 2015 ; Coleman et al, 2021 ). Notably, infection with GB37 yielded delayed cytokine responses at the choriodecidual membranes, despite rapid invasion of the amniotic cavity and neutrophil and CD8+ T recruitment; this is starkly in contrast to the rapid proinflammatory responses associated with infection by hyperpigmented GBS.…”

“…Despite significant immune cell infiltrate to the maternal-fetal interface, digital spatial profiling of the uterine mucosa in GB37-infected NHP revealed that even in tissues where GBS had been detected, inflammation-associated markers were dampened relative to baseline. GB37 also blunted ROS production by neutrophils in a TLR2/4 dependent manner, suggesting that GBS uses HylB to evade host immune recognition by blocking TLR2/4 signaling and downstream ROS production in neutrophils ( Coleman et al, 2021 ). In combination with the prior work on hyperpigmented GBS strains ( Boldenow et al, 2016 ), this study highlights that a chronically catheterized NHP model can demonstrate the diverse roles of various GBS virulence factors in disease pathogenesis that would otherwise be difficult to discern in other animal models.…”