Maternal and neonatal clinical data
Demographic and clinical characteristics of the women and newborns were provided in Table 1. A total of 27 healthy and asymptomatic women were recruited into this study. All of the subjects were Han ethnicity with ages ranging from 19 to 34 years old (mean ages, 28.1 years). The average BMI was 25.2 (range 21.1–34.1). All women gave birth between 37th and 42th gestational week with 14.26 kg (range 9.0–26.30 kg) average gestational weight gain. Most women (21/27) was at labor in 24 hours after hospitalization. The average birth weight of 27 newborns was 3205.9 g (range 2600–3850 g), including 15 boys and 12 girls.
Community structure of vaginal and oral microbiome
To compare the overall vaginal community structures before disinfection and after disinfection, nonmetric multidimensional scaling (NMDS) analysis was implemented on the bacterial abundances. Figure 1A revealed that vaginal samples before disinfection (group BD) separated well from the subjects after disinfection (group AD, p < 0.01, PERMANOVA analysis), whereas vaginal samples after disinfection overlapped with neonatal oral samples (group NO).
Moreover, the vaginal microbiota after disinfection and neonatal oral microbiota was associated with a higher observed species index, evenness index and alpha diversity (Figure 1B). Compared to the vaginal samples before disinfection, the mean observed OTUs numbers increased significantly in neonatal oral samples (179.89 ± 113.37 versus 13.30 ± 14.83, p < 0.01) and in vaginal samples after disinfection (194.33 ± 85.69 versus 13.30 ± 14.83, p < 0.01) (Figure 1B (a)). Accompanied with significantly increased mean pielou index value of group AD (0.55 ± 0.16, p < 0.01) and NO (0.56 ± 0.24, p < 0.01) versus 0.25 ± 0.22 of group BD (Figure 1B (b)). Notably, the mean Shannon index value was 0.92 ± 1.06 of BD group, significantly lower than that of group AD (4.22 ± 1.50, p < 0.01) and NO (4.19 ± 2.22, p < 0.01) (Figure 1B (c)). Consistent with these observations, indices of alpha-diversity and richness were the smallest in samples obtained during pregnancy, with a significant increase in diversity detected in vaginal samples after disinfection and in neonatal oral samples. Further, the weighted UniFrac value of group BD was 0.35 ± 0.29, significant lower than group AD (0.71 ± 0.29, p < 0.01) and NO (0.69 ± 0.27, p < 0.01), which indicated that vaginal microbial communities were more similar within each other in group BD than AD (Figure 1B (d)).
Vaginal and oral microbiome profiling
Microbiome of study participants was characterized using high-throughput sequencing of the 16S rRNA high-variable regions. A total of 3,788,402 reads were included in the analysis. The average sequence read count was 46,770 per sample, with a median of 46,158 (range 13,434–71,182), and the mean and median read lengths were 420 and 423 bp, respectively. The relative abundance of each participant at phylum and genus levels was showed in Figure 2. It showed that the top 10 phyla were Acidobacteria, Actinobacteria, Bacteroidetes, Chloroflexi, Cyanobacteria, Firmicutes, OD1, Planctomycetes, Proteobacteria and Tenericutes (Figure 2A). Meanwhile, the top 10 genera were Atopobium, Enterococcus, Gardnerella, Lactobacillus, Prevotella, Pseudomonas, Ralstonia, Staphylococcus, Streptococcus and Thiobacillus (Figure 2B).
The results also demonstrated that pregnancy was associated with a microbiome largely dominated by phyla Firmicutes (78.65% ± 35.50%) and Actinobacteria (19.83% ± 35.42%), they accounted averagely for more than 98% of the microbial population (Figure 3A). While significant shifts in bacterial phylum structure were observed at neonatal mucous and vaginal samples after disinfection (Figure 3A), with significantly decreased proportions of Firmicutes (37.20% ± 31.99% and 36.75% ± 31.15%) and Actinobacteria (5.44% ± 8.0% and 7.92% ± 15.32%), significantly increased abundance of Proteobacteria (41.94% ± 27.66% and 40.93% ± 26.58% versus 0.35% ± 0.77%) and Bacteroidetes (6.64% ± 8.04% and 4.52% ± 3.97% versus 0.97% ± 2.03%). The differences at the phylum level among the three groups all presented in Figure 3B.
A similar pattern was observed at the genus level (Figure 3C). The mean proportion of Lactobacillus decreased from 73.71% (SD, 38.0%) during pregnancy to 26.42% (SD, 29.88%) after disinfection and 23.94% (SD, 33.03%) in neonatal oral. Similarly, genus Gardnerella accounted averagely 16.43% (SD, 32.24%) before disinfection, decreased to 5.26% (SD, 15.58%) after disinfection and 2.04% (SD, 6.88%) of neonatal oral microbiome. In addition, compared to the pregnancy phase, disinfection phase was also accompanied by increases in genera Streptococcus (3.04% ± 14.68% versus 1.56% ± 6.27%), Ralstonia (16.19% ± 11.50% versus 0.04% ± 0.07%),Pseudomonas (13.45% ± 16.99% versus 0.09% ± 0.27%) and Thiobacillus (1.74% ± 2.94% versus 0%).
Different genus between vaginal and oral microbiome
Next, the STAMP tool was used to analyze bacterial communities in vaginal samples and to detect potential significant differences in relative abundances of genus. Figure 4A included a list of genera that significantly different between vaginal samples during pregnancy and neonatal oral samples. Among them, genera Acinetobacter, Burkholderia, Delftia, Mesorhizobium, Paracoccus, Ralstonia, Reyranella, Salinispora and Shewanella increased significantly in the neonatal microbiome, whereas genus Lactobacillus decreased significantly. In addition, the greatest differences in genus between vaginal samples before disinfection and after disinfection were presented in Figure 4B. Compared to vaginal samples during pregnancy, the relative abundance of genera Acinetobacter, Bradyrhizobium, Burkholderia, Comamonas, Delftia, Mesorhizobium, Mycobacterium, Ralstonia, Reyranella and Salinispora were higher, while genus Lactobacillus was dramatically lower in vaginal microbiome after disinfection. Notably, there was no difference between vaginal samples after disinfection and neonatal oral samples. Collectively, these observations suggested that the microbial composition of the vaginal samples significantly differed between pregnancy and after disinfection according to the relative abundance of sequences.
Community state types analysis and its changes after delivery
Categorizing microbiome profiles based on the taxon with the largest proportion of reads, and hierarchical clustering analysis of bacterial species from the pregnant vaginal microbiome profiles revealed 3 major community state types (CSTs), which showed in Figure 5A. Among all samples, 8 samples were dominated by species L. iners (CST III). Six samples were assigned to CST IV, which were dominated by genus Gardnerella, and also typified by higher proportions of Aerococcus, Atopobium, Bifidobacterium, Corynebacterium, Dialister, Finegoldia, Megasphaera, Mobiluncus, Peptoniphilus, Prevotella, Ralstonia, Staphylococcus, Streptococcus and Sneathia than other CSTs. The rest 13 samples were dominated by species L. helveticus or L. delbrueckii, which were clustered into a new observed community type and names as CST VI in this study.
Dominance of CST IV was observed in neonatal sample and postpartum vaginal samples (n=20 and n=17 in group NO and AD respectively) (Figure 5B). Most subjects of group BD belonged to CST III and CST VI (15/21) switched to CST IV of group NO. Similarly, they (11/21) shifted towards CST IV in vaginal microbiome after disinfection. An interesting change pattern was observed in sample S6, it changed from CST IV to CST VI in the neonatal oral microbiome.
Table 1
Demographic and clinical data for study participants
Characteristics | Value |
Maternal conditions Mother’s age, yrs mean (SD) | 28.1 ± 3.3 |
Mother’s BMI, kg mean (SD) | 25.2 ± 2.6 |
Gestational weight gain, kg mean (SD) | 14.26 ± 0.37 |
Gestational week, wks mean (SD) | 39.5 ± 1.0 |
Hospital stay before labor, hours mean (SD) | 16.8 ± 14.8 |
First degree of perineal laceration, n (%) | 18 (66.7%) |
Neonatal conditions | |
Children sex (male), n (%) | 15 (55.6%) |
Birth weight, g mean (SD) | 3205.9 ± 289.2 |
Apgar score, mean | 10 |