RESUMO
A microbiota vaginal é dominada pelo gênero Lactobacillus, mantendo um pH ácido crucial para a saúde. Alterações hormonais e a menopausa podem impactar essa microbiota. Estreptococos do Grupo B (EGB) são associados a infecções neonatais e podem colonizar a microbiota vaginal. A interação entre Lactobacillus e EGB é complexa, com resultados conflitantes em estudos in vitro e in vivo. O uso de probióticos contendo Lactobacillus pode ter benefícios, como alterar a positividade para EGB em um grupo que recebeu o tratamento. Contudo a replicabilidade deste resultado é limitada, e os mecanismos envolvidos nessa interação ainda são pouco elucidados. Portanto o objetivo desse estudo foi caracterizar cepas de EGB, avaliar sua interação com L. crispatus em diversas condições. Neste estudo foram selecionadas seis cepas, pertencentes aos sorotipos Ia, II, III e V. Essas cepas foram caracterizadas de acordo com o sorotipo capsular, presença de genes de virulência (hialuronidase, ß-citolisina/hemolisina, e ilhas de pili 1, 2a e 2b), resistência a antimicrobianos (penicilina, cefepima, vancomicina, eritromicina e clindamicina), curva de crescimento, Restriction Fragment Length Polymorphism (RFLP) e Multi-Locus Sequence Typing (MLST). O resultado de MLST identificou os sequence types ST1, ST23 ST28 e ST182, e os clonal complexes CC1, CC19 e CC23. Cepas possuem a maior parte dos genes de virulência testados. Somente uma cepa apresentou alguma resistência, no caso à eritromicina e clindamicina. Resultados que estão de acordo com a literatura como os principais descritos em isolados clínicos. Os ensaios de interação realizados foram: crescimento em co-cultura, formação de biofilme, ensaios de adesão e exclusão em cultura celular e por fim, análise do perfil proteico. Alguns efeitos de L. crispatus no EGB observados foram aumento na formação de biofilme de EGB e diminuição da sua proliferação quando em co-cultura. Contudo esses resultados variaram de acordo com a cepa de EGB. Nos ensaios de competição em cultura celular, a presença de L. crispatus levou à diminuição da adesão em uma cepa pertencente ao sorotipo III, e aumento de adesão em uma cepa pertencente ao sorotipo V. Quando a cultura de células recebeu tratamento prévio de L. crispatus, nos ensaios de exclusão, houve menor adesão de EGB. Contudo todos os efeitos observados são cepa-dependente; não foram encontradas associações entre o comportamento das cepas e seu sorotipo capsular ou fenótipo MLST. Estudos futuros podem elucidar as respostas ativadas na presença de microrganismos comensais
The vaginal microbiota is dominated by the genus Lactobacillus, which helps to maintain a crucial acidic pH for health. Hormonal changes and menopause can impact this microbiota. Group B Streptococcus (GBS) are associated with neonatal infections and can colonize the vaginal microbiota. The interaction between Lactobacillus and GBS is complex, with conflicting results in in vitro and in vivo studies. The use of probiotics containing Lactobacillus may have benefits, such as altering GBS positivity in a treated group. However, the replicability of this result is limited, and the mechanisms involved in this interaction are still poorly understood. The use of probiotics containing Lactobacillus may have some benefits, but further research is needed. These strains were characterized based on capsular serotype, presence of virulence genes (hyaluronidase, ß-cytolysin/hemolysin, and pili islands 1, 2a, and 2b), antimicrobial resistance (penicillin, cefepime, vancomycin, erythromycin, and clindamycin), growth curve, Restriction Fragment Length Polymorphism (RFLP), and Multi-Locus Sequence Typing (MLST) Therefore, the aim of this study was to characterize GBS strains, evaluate their interaction with L. crispatus under various conditions. Six strains were selected for this study, belonging to serotypes Ia, II, III, and V. MLST results identified sequence types ST1, ST23, ST28, and ST182, and clonal complexes CC1, CC19, and CC23. Most strains possessed the tested virulence genes, with only one strain showing resistance to erythromycin and clindamycin. These results are in line with the literature. Interaction assays included co-culture growth, biofilm formation, adhesion and exclusion assays in cell culture, and, finally, protein profile analysis. Some observed effects of L. crispatus on GBS included an increase in GBS biofilm formation and a decrease in GBS proliferation during co-culture. However, these results varied according to the GBS strain. GBS interaction with L. crispatus resulted in increased biofilm formation and decreased proliferation in co-culture. In cell culture competition assays, the presence of L. crispatus led to decreased adhesion in one GBS strain belonging to serotype III and increased adhesion in one strain belonging to serotype V. When cell culture received pre-treatment with L. crispatus, exclusion assays showed lower GBS adhesion. However, all observed effects are strain-dependent; no associations were found between strain behavior and capsular serotype or MLST phenotype. Future studies may elucidate the responses activated in the presence of commensal microorganisms
Assuntos
Streptococcus agalactiae/classificação , Lactobacillus crispatus/classificação , Técnicas In Vitro/métodos , Técnicas de Cultura de Células/métodos , Infecções/complicaçõesRESUMO
The microbial composition of the human vagina differs from that of all other mammals, likely as a consequence of the unique composition of vaginal constituents that promote the selective maintenance and proliferation of distinct bacterial species. Similarly, variations between individual healthy women in genetic, environmental, and medical variables also modify the vaginal lumen composition. The direction and magnitude of immune reactions to microorganisms present in the vagina, responses to stress and non-infectious stimuli, coupled with medical and pregnancy history and environmental exposures, can greatly differ between women. Adaptations to both internal and external pressures will determine the ability of select resident vaginal bacteria to numerically dominate and, therefore, the definition of a "normal" vaginal microbiota will substantially differ between individual healthy women.
Assuntos
Microbiota , Vagina , Gravidez , Animais , Feminino , Humanos , Vagina/microbiologia , Microbiota/fisiologia , Reprodução , Bactérias , RNA Ribossômico 16S , Mamíferos/genéticaRESUMO
Background: Intra-continentally, vaginal microbiome signatures are reported to be significantly different between Black and Caucasian women, with women of African ancestry having the less well defined heterogenous bacterial community state type (CST) deficient of Lactobacillus species (CST IV). The objective of this study was to characterize the vaginal microbiomes across a more diverse intercontinental group of women (N = 151) of different ethnicities (African American, African Kenyan, Afro-Caribbean, Asian Indonesian and Caucasian German) using 16S rRNA gene sequence analysis to determine their structures and offer a comprehensive description of the non-Lactobacillus dominant CSTs and subtypes. Results: In this study, the bacterial composition of the vaginal microbiomes differed significantly among the ethnic groups. Lactobacillus spp. (L. crispatus and L. iners) dominated the vaginal microbiomes in African American women (91.8%) compared to European (German, 42.4%), Asian (Indonesian, 45.0%), African (Kenyan, 34.4%) and Afro-Caribbean (26.1%) women. Expanding on CST classification, three subtypes of CST IV (CST IV-A, IV-B and IV-C) (N = 56, 37.1%) and four additional CSTs were described: CST VI Gardnerella vaginalis-dominant (N = 6, 21.8%); CST VII (Prevotella-dominant, N = 1, 0.66%); CST VIII (N = 9, 5.96%), resembling aerobic vaginitis, was differentiated by a high proportion of taxa such as Enterococcus, Streptococcus and Staphylococcus (relative abundance [RA] > 50%) and CST IX (N = 7, 4.64%) dominated by genera other than Lactobacillus, Gardnerella or Prevotella (e.g., Bifidobacterium breve and Anaerococcus vaginalis). Within the vaginal microbiomes, 32 "taxa with high pathogenic potential" (THPP) were identified. Collectively, THPP (mean RA ~5.24%) negatively correlated (rs = -0.68, p < 2.2e-16) with Lactobacillus species but not significantly with Gardnerella/Prevotella spp. combined (r = -0.13, p = 0.1). However, at the individual level, Mycoplasma hominis exhibited moderate positive correlations with Gardnerella (r = 0.46, p = 2.6e-09) and Prevotella spp. (r = 0.47, p = 1.4e-09). Conclusions: These findings while supporting the idea that vaginal microbiomes vary with ethnicity, also suggest that CSTs are more wide-ranging and not exclusive to any particular ethnic group. This study offers additional insight into the structure of the vaginal microbiome and contributes to the description and subcategorization of non-Lactobacillus-dominated CSTs.
Assuntos
Microbiota , Vagina , Feminino , Humanos , Masculino , RNA Ribossômico 16S/genética , Quênia , Vagina/microbiologia , Microbiota/genética , Lactobacillus/genética , Bactérias/genética , Gardnerella/genéticaRESUMO
BACKGROUND: Interplay between vaginal microbiome and human papillomavirus (HPV) remains unclear, partly due to heterogeneity of microbiota. METHODS: We used data from 546 women enrolled in a cross-sectional study in 5 Brazil. We genotyped vaginal samples for HPV and sequenced V3-V4 region of 16S rRNA gene for vaginal microbiome analysis. We used stepwise logistic regression to construct 2 linear scores to predict high-risk HPV (hrHPV) positivity: one based exclusively on presence of individual bacterial taxa (microbiome-based [MB] score) and the other exclusively on participants' sociodemographic, behavioral, and clinical (SBC) characteristics. MB score combined coefficients of 30 (of 116) species. SBC score retained 6 of 25 candidate variables. We constructed receiver operating characteristic curves for scores as hrHPV correlates and compared areas under the curve (AUC) and 95% confidence intervals (CI). RESULTS: Overall, prevalence of hrHPV was 15.8%, and 26.2% had a Lactobacillus-depleted microbiome. AUCs were 0.8022 (95% CI, .7517-.8527) for MB score and 0.7027 (95% CI, .6419-.7636) for SBC score (Pâ =â .0163). CONCLUSIONS: The proposed MB score is strongly correlated with hrHPV positivity-exceeding the predictive value of behavioral variables-suggesting its potential as an indicator of infection and possible value for clinical risk stratification.
Assuntos
Alphapapillomavirus , Microbiota , Infecções por Papillomavirus , Neoplasias do Colo do Útero , Alphapapillomavirus/genética , Estudos Transversais , Feminino , Humanos , Microbiota/genética , Papillomaviridae/genética , Infecções por Papillomavirus/epidemiologia , RNA Ribossômico 16S/genética , Vagina/microbiologiaRESUMO
PURPOSE: To investigate associations between bacterial species in the vagina in mid-trimester pregnant women from Brazil. METHODS: The vaginal microbiome in 613 subjects was identified by analysis of the V1-V3 region of bacterial 16S ribosomal RNA and the relative prevalence of individual bacteria were determined. RESULTS: The bacterial species present in the greatest number of women were Lactobacillus crispatus (306 women), L. iners (298 women) and Gardnerella vaginalis (179 women). When present in the vagina, L. crispatus was the most abundant bacterium more than 85% of the time. In contrast, L. iners and G. vaginalis were most abundant in 63% and 41% of women who were positive for these microorganisms, respectively (p < 0.0001 vs. L. crispatus). The proportion of L. crispatus was negatively associated with the proportions of L. iners, L. jensenii, L. gasseri, G. vaginalis, Megasphaera, Atopobium vaginae and Prevotella (p < 0.0001). In contrast, the proportion of G. vaginalis was positively associated with levels of Megasphaera, A. vaginae and Prevotella (p < 0.0001) while L. iners proportion was unrelated to the proportion of L. jensenii, G. vaginalis, Megasphaera, A. vaginae or Prevotella. CONCLUSION: The composition of the vaginal microbiota in mid-trimester pregnant women is influenced by the relative concentrations of L. crispatus, L. iners and G. vaginalis.
Assuntos
Gardnerella vaginalis/isolamento & purificação , Lactobacillus crispatus/isolamento & purificação , Lactobacillus/isolamento & purificação , Reação em Cadeia da Polimerase/métodos , Vagina/microbiologia , Vaginose Bacteriana/microbiologia , Adulto , Bactérias , Brasil/epidemiologia , Feminino , Humanos , Gravidez , Segundo Trimestre da Gravidez , Gestantes , Prevalência , Estudos Prospectivos , RNA Ribossômico 16S/genética , Vaginose Bacteriana/epidemiologiaRESUMO
OBJECTIVE: To determine the presence and identity of extracellular bacteriophage (phage) families, genera and species in the vagina of pregnant women. DESIGN: Descriptive, observational cohort study. SETTING: São Paulo, Brazil. POPULATION: Pregnant women at 21-24 weeks' gestation. METHODS: Vaginal samples from 107 women whose vaginal microbiome and pregnancy outcomes were previously determined were analysed for phages by metagenomic sequencing. MAIN OUTCOME MEASURES: Identification of phage families, genera and species. RESULTS: Phages were detected in 96 (89.7%) of the samples. Six different phage families were identified: Siphoviridae in 69.2%, Myoviridae in 49.5%, Microviridae in 37.4%, Podoviridae in 20.6%, Herelleviridae in 10.3% and Inviridae in 1.9% of the women. Four different phage families were present in 14 women (13.1%), three families in 20 women (18.7%), two families in 31 women (29.1%) and one family in 31 women (29.1%). The most common phage species detected were Bacillus phages in 48 (43.6%), Escherichia phages in 45 (40.9%), Staphylococcus phages in 40 (36.4%), Gokushovirus in 33 (30.0%) and Lactobacillus phages in 29 (26.4%) women. In a preliminary exploratory analysis, there were no associations between a particular phage family, the number of phage families present in the vagina or any particular phage species and either gestational age at delivery or the bacterial community state type present in the vagina. CONCLUSIONS: Multiple phages are present in the vagina of most mid-trimester pregnant women. TWEETABLE ABSTRACT: Bacteriophages are present in the vagina of most pregnant women.
Assuntos
Bacteriófagos , Microbiota/fisiologia , Vagina/microbiologia , Adulto , Bacteriófagos/classificação , Bacteriófagos/genética , Bacteriófagos/isolamento & purificação , Brasil , Feminino , Idade Gestacional , Humanos , Metagenoma , Metagenômica/métodos , Metagenômica/estatística & dados numéricos , Gravidez , Resultado da Gravidez/epidemiologiaRESUMO
OBJECTIVES: Short cervical length is a predictor of preterm birth. We evaluated if there were racial differences in variables associated with cervical length in pregnant Brazilian women. METHODS: Cervical length was determined by vaginal ultrasound in 414 women at 21 weeks gestation. All women were seen at the same clinic and analyzed by the same investigators. Women found to have a short cervix (≤25 mm) received vaginal progesterone throughout gestation. Composition of the vaginal microbiome was determined by analysis of the V1-V3 region of the gene coding for bacterial 16S ribosomal RNA. Demographic, clinical and outcome variables were determined by chart review. Subjects were 53.4% White, 37.2% mixed race and 9.4% Black. RESULTS: Pregnancy, medical history and education level were similar in all groups. Mean cervical length was shorter in Black women (28.4 mm) than in White (32.4 mm) or mixed race (32.8 mm) women (p≤0.016) as was the percentage of women with a short cervix (23.1, 12.2, 7.8% in Black, White, mixed race respectively) (p≤0.026). Mean cervical length increased with maternal age in White (p=0.001) and mixed race (p=0.045) women but not Black women. There were no differences in bacterial dominance in the vaginal microbiota between groups. Most women with a short cervix delivered at term. CONCLUSIONS: We conclude that Black women in Brazil have a shorter cervical length than White or mixed race women independent of maternal age, pregnancy and demographic history or composition of the vaginal microbiome.
Assuntos
Medida do Comprimento Cervical , Colo do Útero/diagnóstico por imagem , Nascimento Prematuro , Ultrassonografia Pré-Natal , Adulto , Brasil/epidemiologia , Medida do Comprimento Cervical/métodos , Medida do Comprimento Cervical/estatística & dados numéricos , Feminino , Idade Gestacional , Humanos , Idade Materna , Microbiota/genética , Gravidez , Resultado da Gravidez/epidemiologia , Nascimento Prematuro/diagnóstico , Nascimento Prematuro/etnologia , Nascimento Prematuro/fisiopatologia , Nascimento Prematuro/prevenção & controle , RNA Ribossômico 16S/isolamento & purificação , Fatores Raciais , Ultrassonografia Pré-Natal/métodos , Ultrassonografia Pré-Natal/estatística & dados numéricos , Vagina/microbiologiaRESUMO
Vaginal samples from women with term deliveries were tested for torquetenovirus (TTV) by gene amplification, matrix metalloproteinase (MMP)-8 and D- and L-lactic acid by ELISA, and microbiome composition by analysis of the bacterial 16S ribosomal RNA gene. TTV was detected in 43.2%, 31.5%, and 41.4% of first trimester, third trimester, and postpartum samples, respectively. The viral titer was higher in postpartum than in the first (p = 0.0018) or third (p = 0.0013) trimester. The mean gestational age at delivery was lower in women positive for TTV in their first trimester (p = 0.0358). In the first and third trimester, the MMP-8 level was higher if TTV was also present (p < 0.0091). The D-lactic acid level was lower in first trimester samples if TTV was present (p = 0.0334). Lactobacillus crispatus dominance in first and third trimester samples was higher when TTV was absent (p < 0.0033). We conclude that TTV is present in the vagina in many women with normal pregnancy outcomes and that its occurrence is associated with a lack of L. crispatus dominance, an increase in vaginal MMP-8 and a decrease in D-lactic acid.
Assuntos
Infecções por Vírus de DNA , Ácido Láctico/análise , Lactobacillus crispatus , Metaloproteinase 8 da Matriz/análise , Complicações na Gravidez/virologia , Torque teno virus , Vagina/virologia , Adulto , Líquidos Corporais/virologia , Feminino , Humanos , Lactobacillus crispatus/isolamento & purificação , Período Pós-Parto , Gravidez , Resultado da Gravidez , Trimestres da Gravidez , Torque teno virus/isolamento & purificaçãoRESUMO
Preterm birth remains the main contributor to early childhood mortality. The vaginal environment, including microbiota composition, might contribute to the risk of preterm delivery. Alterations in the vaginal microbial community structure might represent a risk factor for preterm birth. Here, we aimed to (a) investigate the association between preterm birth and the vaginal microbial community and (b) identify microbial biomarkers for risk of preterm birth. Microbial DNA was isolated from vaginal swabs in a cohort of 69 women enrolled at hospital admission for their delivery. Microbiota was analyzed by high-throughput 16S rRNA sequencing. While no differences in microbial diversity measures appeared associated with the spontaneous preterm and full-term outcomes, the microbial composition was distinct for these groups. Differential abundance analysis showed Lactobacillus species to be associated with full-term birth whereas an unknown Prevotella species was more abundant in the spontaneous preterm group. Although we studied a very miscegenated population from Brazil, our findings were similar to evidence pointed by other studies in different countries. The role of Lactobacillus species as a protector in the vaginal microbiome is demonstrated to be also a protector of spontaneous preterm outcome whereas the presence of pathogenic species, such as Prevotella spp., is endorsed as a factor of risk for spontaneous preterm delivery.
Assuntos
Bactérias/classificação , Microbiota , Trabalho de Parto Prematuro/diagnóstico , Vagina/microbiologia , Adulto , Bactérias/isolamento & purificação , Biomarcadores/análise , Brasil , Estudos de Coortes , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lactobacillus/classificação , Trabalho de Parto Prematuro/microbiologia , Gravidez , Prevotella/classificação , Prevotella/patogenicidade , RNA Ribossômico 16S , Fatores de Risco , Adulto JovemRESUMO
The female lower genital tract harbors a complex microbial community essential for homeostasis and health. During pregnancy, the female body undergoes unique hormonal changes that contribute to weight gain as well as modulations in immune function that can affect microbiota composition. Several studies have described the vaginal microbiota of pregnant women from the USA, Europe and Mexico. Here we expand our knowledge about the vaginal microbial communities during the third trimester to healthy expectant Brazilian mothers. Vaginal samples were collected from patients delivering at the Hospital de Clínicas de Porto Alegre, Brazil. Microbial DNA was isolated from samples and the V4 region of the 16S rRNA gene was amplified and sequenced using the PGM Ion Torrent. Brazilian pregnant women presented three distinct types of microbial community at the time of labor. Two microbial communities, Cluster 1 and Cluster 3, presented an overall dominance of Lactobacillus while Cluster 2 tended to present higher diversity and richness, with the presence of Pseudomonas, Prevotella and other vaginosis related bacteria. About half of the Brazilian mothers sampled here had dominance of L. iners. The proportion of mothers without dominance of any Lactobacillus was higher in Brazil (22%) compared to UK (2.4%) and USA, where this community type was not detected. The vaginal microbiota showed significant correlation with the composition of the babies' gut microbiota (p-value = 0.002 with a R2 of 15.8%). Mothers presenting different vaginal microbiota shared different microorganisms with their newborns, which would reflect on initial colonizers of the developing newborns' gut.