Objectives To study Group B Streptococcus (GBS) isolates associated with different clinical syndromes: asymptomatic carriage in pregnant women, intrauterine fetal death (IUFD), and early onset disease (EOD) in the newborn. Methods GBS isolates were collected from asymptomatic pregnant women admitted for labor, IUFD cases, and neonates with EOD. Serotypes and antibiotic susceptibilities were determined. Multilocus sequence typing (MLST) was performed to assess genetic epidemiology. Results GBS carriage rate was 26.1% (280/1074). The dominant serotype among asymptomatic pregnant women was VI [98/240 women (40.8%)], followed by serotypes III, V and IV in 42/240 (17.5%), 30/240 (12.5%) and 28/240 (11.7%) women, respectively. The dominant serotype in IUFD cases was serotype VI [10/13 (76.9%)]. In contrast the prevalent serotype among EOD cases was III [16/19 (84.2%)]. ST-1 was associated with IUFD [7/13 (53.8%)], ST-17 was associated with serotype III and EOD in the newborn 14/19 (73.7%)]. Erythromycin and clindamycin resistance reached 36.8%, 7.7% and 20.0%among EOD, vaginal carriage and IUFD, respectively. Conclusions Serotypes VI and ST-1 were dominant among asymptomatic pregnant women and in IUFD cases while EOD was associated with serotype III and ST-17. Invasive mechanisms thus may differ between IUFD and EOD in the newborn and virulence may be related to capsule serotype. Resistance rates to erythromycin and clindamycin were high in EOD cases.

BackgroundGroup B streptococcus (GBS) harbors many virulence factors but there is limited data regarding their importance in colonization in pregnancy and early-onset disease (EOD) in the newborn. We hypothesized that colonization and EOD are associated with different distribution and expression of virulence factors.MethodsWe studied 36 GBS EOD and 234 GBS isolates collected during routine screening. Virulence genes (pilus-like structures-PI-1, PI-2a, PI-2b; rib and hvgA) presence and expression were identified by PCR and qRT-PCR. Whole genome sequencing (WGS) and comparative genomic analyses were used to compare coding sequences (CDSs) of colonizing and EOD isolates.ResultsSerotype III (ST17) was significantly associated with EOD and serotype VI (ST1) with colonization. hvgA and rib genes were more prevalent among EOD isolates (58.3 and 77.8%, respectively; p < 0.01). The pilus loci PI-2b and PI-2a were more prevalent among EOD isolates (61.1%, p < 0.01), while the pilus loci PI-2a and PI-1 among colonizing isolates (89.7 and 93.1% vs. 55.6 and 69.4%, p < 0.01). qRT PCR analysis revealed that hvgA was barely expressed in colonizing isolates, even though the gene was detected. Expression of the rib gene and PI-2b was two-fold higher in EOD isolates compared to colonizing isolates. Transcription of PI-2a was three-fold higher in colonizing isolates compared to EOD isolates. ST17 isolates (associated with EOD) had a smaller genome size compared ST1 and the genome was more conserved relative to the reference strain and ST17 isolates. In a multivariate logistic regression analysis virulence factors independently associated with EOD were serotype 3, and PI-1 and PI-2a was protective.ConclusionThere was a significant difference in the distribution of hvg A, rib, and PI genes among EOD (serotype III/ST17) and colonizing (serotype VI/ST1) isolates suggesting an association between invasive disease and these virulence factors. Further study is needed to understand the contribution of these genes to GBS virulence.

IntroductionGBS may cause a devastating disease in newborns. In early onset disease of the newborn the bacteria are acquired from the colonized mother during delivery. We characterized type VII secretion system (T7SS), exporting small proteins of the WXG100 superfamily, in group B Streptococci (GBS) isolates from pregnant colonized women and newborns with early onset disease (EOD) to better understand T7SS contribution to virulence in these different clinical scenarios.MethodsGBS genomes [N=33, 17 EOD isolates (serotype III/ST17) and 16 colonizing isolates (12 serotype VI/ST1, one serotype VI/ST19, one serotype VI/ST6, and two serotype 3/ST19)] were analyzed for presence of T7SS genes and genes encoding WXG100 proteins. We also perform bioinformatic analysis. Galleria mellonella larvae were used to compare virulence between colonizing, EOD, and mutant EOD isolates. The EOD isolate number 118659 (III/ST17) was used for knocking out the essC gene encoding a membrane-bound ATPase, considered the driver of T7SS.ResultsMost GBS T7SS loci encoded core component genes: essC, membrane-embedded proteins (essA; essB), modulators of T7SS activity (esaA; esaB; esaC) and effectors: [esxA (SAG1039); esxB (SAG1030)].Bioinformatic analysis indicated that based on sequence type (ST) the clinicalGBS isolates encode at least three distinct subtypes of T7SS machinery. In all ST1isolates we identified two copies of esxA gene (encoding putative WXG100proteins), when only 23.5% of the ST17 isolates harbored the esxA gene. Five ST17isolates encoded two copies of the essC gene. Orphaned WXG100 molecule(SAG0230), distinct from T7SS locus, were found in all tested strains, except inST17 strains where the locus was found in only 23.5% of the isolates. In ST6 andST19 isolates most of the structure T7SS genes were missing. EOD isolates demonstrated enhanced virulence in G. mellonella modelcompared to colonizing isolates. The 118659DessC strain was attenuated in itskilling ability, and the larvae were more effective in eradicating 118659DessC.ConclusionsWe demonstrated that T7SS plays a role during infection. Knocking out the essC gene, considered the driver of T7SS, decreased the virulence of ST17 responsible for EOD, causing them to be less virulent comparable to the virulence observed in colonizing isolates.