RESUMEN
The vaginal microbiome plays a critical role in determining the progression of female genital tract infections; however, little is known about the vaginal microbiota of Indian women. We aimed to investigate the vaginal microbial architecture of women with asymptomatic bacterial vaginosis (BV) (n=20) and normal microbiota (n=19). Microbial diversity was analyzed in vaginal swabs from regularly menstruating women (18-45yrs) by 16S rRNA V3-V4 amplicon (MiSeq Illumina) sequencing. Rarefaction analysis showed a higher number of species in normal flora compared to BV. Alpha diversity as measured by Pielou's evenness revealed microbial diversity was significantly greater in BV samples than normal microbiota (p= 0.0165). Beta diversity comparison using UniFrac metrics indicated distinct microbial communities clustering between normal and BV flora. Firmicutes were the major phyla observed in vaginal specimens of normal microbiota whereas Actinobacteria, Fusobacteria, Bacteroidetes were significantly abundant in BV samples. Notably, the relative abundance of Lactobacillus was significantly high in normal microbiota. Conversely Gardnerella, Sneathia, Prevotella, Atopobium, Ureaplasma, Dialister significantly dominated dysbiotic microbiota. Relative frequency of Lactobacillus decreased significantly in BV (6%) as compared to normal microbiota (35.2%). L. fermentum, L. gasseri, L. iners, L. jensenii, L. mucosae, L. ruminis, L. salivarius, L. coleohominis was more exclusively present in normal microbiota. L. iners was detected from both the groups with a relative frequency of 50.4% and 17.2% in normal and BV microbiota respectively. Lefse analysis indicated Atopobium vaginae, Sneathia amnii, Mycoplasma hominis Prevotella disiens in the vaginal microbiota as a biomarker for dysbiosis and L. jensenii as a biomarker of a healthy microbiota. Firmicutes were negatively correlated to Tenericutes, Actinobacteria, Bacteroidetes, and Fusobacteria. Proteobacteria positively correlated to Tenericutes, and Bacteroidetes were shown to be positively correlated to Fusobacteria. Predicted functional analysis indicated differences in the functional profiles between BV and normal microbiota. Normal microbiota utilized pathways essential for phosphatidylglycerol biosynthesis I & II, peptidoglycan biosynthesis, geranylgeranyl diphosphate biosynthesis I, mevalonate pathway, CoA biosynthesis pathway I and pyrimidine nucleotide salvage; whereas BV bacteria had characteristic aromatic amino acid biosynthesis, pentose phosphate pathway, carbohydrate degradation. In conclusion, women with asymptomatic BV have vaginal microbiota significantly different than women with normal microbiota. Furthermore, the study provides insights into the vaginal microbial structure of Indian women that will enable us to explore the prospective candidates for restoring the vaginal microbiota.
Asunto(s)
Disbiosis , Microbiota , Femenino , Humanos , Microbiota/genética , Estudios Prospectivos , ARN Ribosómico 16S/genética , Vagina/microbiologíaRESUMEN
Asymptomatic bacterial vaginosis (BV) in reproductive-age women has serious obstetric and gynecological consequences. Despite its high incidence, the behavior of vaginal lactobacilli in asymptomatic BV is unknown. We analyzed the functional properties of previously isolated vaginal lactobacilli from asymptomatic women with normal, intermediate, and BV microbiota. Lactic acid and antimicrobial activity against seven urogenital pathogens were evaluated from lactobacilli cell-free culture supernatants (CFCs) (n = 207) after 48 h incubation in MRS. Lactobacilli isolates were used to evaluate H2O2, autoaggregation and coaggregation with C. albicans. Lactobacilli from normal microbiota produced more d-lactate than lactobacilli from intermediate and asymptomatic BV (p = 0.007). L. plantarum, L. fermentum and L. reuteri produced greater d-lactate whereas L. rhamnosus, L. crispatus, L. johnsonii were greater producers of l-lactate. Interspecies positive correlation was observed in the lactic acid contents of CFCs. Distribution of H2O2-producing lactobacilli did not vary significantly among the groups. When lactic acid isomers were considered, species from intermediate and BV microbiota clustered together with each other and distinctly from species of normal microbiota. Broad-spectrum antagonism (≥90% inhibition) against E. coli, C. albicans, S. aureus, P. aeruginosa, G. vaginalis, N. gonorrhoeae, S. agalactiae were displayed by 46.86% (97) of isolates. Our study highlights the differential functional properties of vaginal lactobacilli from women with normal microbiota and asymptomatic BV.
RESUMEN
Vaginal microbiota contributes in maintaining and protecting the urogenital niche from infections and their sequelae. Despite extensive research, microbiome studies have often ignored asymptomatic bacterial vaginosis (BV) and vulvovaginal candidiasis (VVC). The present study aimed to explore the cultivable vaginal bacterial and mycological communities in women asymptomatic for BV and VVC using multiplex PCR and species-specific PCR. Vaginal swabs collected from 199 participants asymptomatic for urogenital infections, scored by Nugent criteria indicated 73.9% had normal microbiota, 11.6% intermediate and 14.5% BV. The most frequent Lactobacillus species in normal women were L. iners (69.4%), L. crispatus (24.5%), L. reuteri (20.4%). Women with BV colonized L. iners (62.1%); L. rhamnosus (41.4%); L. salivarius (13.7%) and L. reuteri (7.2%). Furthermore, L. crispatus was associated with normal microbiota, whereas L. iners was a frequent member of normal and dysbiotic microbiota. Lactobacillus abundance and species richness reduced in asymptomatic BV. Also L. crispatus, L. fermentum, L. acidophilus and L. delbruckii were absent in these women. L. iners significantly co-existed with other Lactobacillus species, indicating its failure in independently maintaining the healthy vaginal niche. Of 30.4% women detected with Candida, 72.1% constituted non-albicans Candida. Predominance of C. albicans increased from 18.4% in healthy to 60% in women with asymptomatic BV; whereas distribution of BV related bacteria did not vary across the groups. Heterogeneous population of lactobacilli in 80.8% of normal women calls attention towards cumulative effects of these species in safeguarding the vaginal microenvironment. Since the microbiota of asymptomatic BV was different from healthy, screening and management could be encouraged to avoid further complications of infections.
Asunto(s)
Candidiasis Vulvovaginal/microbiología , Microbiota/fisiología , Vagina/microbiología , Vaginosis Bacteriana/microbiología , Adolescente , Adulto , Biodiversidad , Candida/clasificación , Candida/genética , Candida/crecimiento & desarrollo , Candida/aislamiento & purificación , Candidiasis Vulvovaginal/genética , ADN Bacteriano , ADN de Hongos , Disbiosis , Femenino , Humanos , Lactobacillus/clasificación , Lactobacillus/genética , Lactobacillus/crecimiento & desarrollo , Lactobacillus/aislamiento & purificación , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S , Especificidad de la Especie , Vaginosis Bacteriana/genética , Adulto JovenRESUMEN
INTRODUCTION: Lactobacillus dominated vaginal microenvironment is associated with lower risk of genital infections. Numerous studies have reported geographic and ethnic variations in vaginal microbiome structure between healthy individuals from different race and ethnicity. India has a great diversity, so it is intriguing to find out if such divergences exist in vaginal lactobacilli. The present study aimed to investigate predominant Lactobacillus species in vaginas of healthy Indian women and screen isolates for lactic acid and H2O2 production. METHODOLOGY: 203 premenopausal women asymptomatic for any vaginal complaints were recruited. The lactobacilli isolates on MRS agar were identified by Multiplex-PCR and 16sRNA gene sequencing. RAPD was used to differentiate strains of same species. H2O2 and lactic acid was evaluated on TMB-HRP MRS agar and BCP-MRS agar respectively. RESULTS: Lactobacilli were recovered from 107/109 (98.2%) women with normal microflora. L. iners 64.7% (68), L. crispatus 26.7% (28), L. reuteri 21.9% (23), L. jensenii 16.2% (17) and L. gasseri 15.2% (16) were the most frequently occurring vaginal lactobacilli in normal women. The vaginal microflora was dominated by either by a single (80%, n = 84) or a combination (20%, n = 21) of Lactobacillus species. Though most frequently identified, L. iners, coexisted only with other Lactobacillus species. All isolates were acid producers but H2O2 was produced by 94.2% isolates. CONCLUSIONS: Our study reports prevalent vaginal lactobacilli which could be explored as probiotics. Presence of heterogeneous Lactobacillus population highlights the cumulative effects of different lactobacilli maintaining vaginal health. Contrasting observations about L. iners reiterates its puzzling role in vaginal immunity, advocating further research.