1)Vaginal microbiome of reproductive-age women

vaginal microbiomes help prevent urogenital diseases in women and maintain health.
396 asymptomatic North American women Ethnicity-white, black, Hispanic, and Asian

species composition characterized by pyrosequencing of barcoded 16S rRNA genes.

The communities clustered into five groups: four

were dominated by Lactobacillus iners, L. crispatus, L. gasseri, or

L. jensenii, whereas the fifth had lower proportions of lactic acid

bacteriaFacultative Anaerobes Mobiluncus, and Finegoldia Atopobium, Corynebacterium, Anaerococcus,

Peptoniphilus, Prevotella, Gardnerella, Sneathia, Eggerthella, , among

(CST IV-A and IV-B). CST IV-A comprised a variety of anaerobic bacteria (including Streptococcus and
Prevotella),
whereas CST IV-B harbored higher proportions of the genera Atopobium, Gardnerella, and Prevotella among
others.

address three aims.

The first was to establish whether there were correlations between community composition and vaginal pH because these would be indicative of
community performance.

The second was to explore how the species composition of vagi-nal communities was reflected in Nugent scores (25), a diagnosticfactor
commonly used to identify women with bacterial vaginosis(26).

Finally, the third aim was to identify patterns in the relative of different species because these might reflect antag-

onistic or cooperative interspecies interactions.

Each woman used two swabs to self-collect midvaginal samples. One was used to evaluate the vaginal microbiota on the basis of the Nugent
criteria used for the diagnosis of bacterial vaginosis and the second was used in procedures to determine the composition and structure of the
resident bacterial communities (27)

The human vaginal microbiota seem to play a key role in preventing a number of urogenital diseases, such as bacterial vaginosis, yeast infections,
sexually transmitted infections, urinary tract infections (2–9), and HIV infection (10, Common wisdom attributes this to lactic acid–producing
bacteria, mainly Lactobacillus sp., that commonly inhabit the vagina. These species are thought to play key protective roles by lowering the
environmental pH through lactic acid production (12, 13), by producing various bacteriostatic and bacteriocidal compounds,

If a vaginal bacterial community has ≈108 cells per milliliter of vaginal secretion, then high numbers (105 cells/mL) of “rare” members are
present in the community, phylotypes present at densities of <105 cells per milli-liter would remain undetected.

Communities in group I, which occurred in 26.2% of the women sampled, were dominated by L. crispatus, whereas groups II (6.3%) domi-nated
by L. gasseri, III (34.1%,. L iners), and V (5.3%) were domi-nated, L. jensenii

The Nugent criteria(25) are widely used to diagnose bacterial vaginosis according to the proportions of different cellular morphologies seen in
gram stained smears of vaginal samples

Sained smears of vaginal samples. The weighted score computed using these criteria is thought to reflect the relative abundance of the following
morphotypes: lactobacilli, Gardnerella vaginalis or Bacteroides (small gram-variable rods or Gram-negative rods), and curved gram-variable
rods. The resulting scores range from 0 to 10, with those of 7 and higher considered to be indicative of bacterial vaginosis, whereas scores of 4–6
and 3 or less are considered intermediate and normal, respectively. In this study communities with high Nugent scores were most often associated
with communities in group IV but were also observed in com-munities belonging to other groups (Fig. 1).

Vaginal bacterial communities dominated by species of Lactobacillus

(groups I, II, III, and V) were found in 80.2% and89.7% of Asian and white women, respectively, but in only 59.6%and 61.9% of Hispanic and
black women, respectively. Thehigher median pH values in Hispanic (pH 5.0 ± 0.59) and black (pH 4.7 ± 1.04) women reflects the higher
prevalence of communities not dominated by Lactobacillus sp. (cluster IV) in these two ethnic groups when compared with Asian (pH 4.4 ±
0.59)and white (pH 4.2 ± 0.3) women (Table 1). This is significant because the occurrence of high numbers of lactobacilli and pH<4.5 have
become synonymous with“healthy”. If
We found that community group IV (diverse group) was overrepresented in Hispanic (34.3%) and black (38.9%) women as compared with Asian
(17.6%) and white (9.3%) women (

high Nugent scores, Aerococcus, Anaeroglobus,Anaerotruncus, Atopobium, Coriobacteriaceae_2, Dialister, Eggerthella,

Gardnerella, Gemella, Megasphaera, Mobiluncus, Parvi monas,
Peptoiphilus, Prevotella, Porphyomonas, Pr evotellaceae_1,Prevotellaceae_2, Ruminococcaceae, and Snethia

2)Cervicovaginal microbiota, women's health, and reproductive outcomes

Lactobacillus spp. Dominated vaginal microbiota have been intrinsi-cally linked to estrogen production
and the accumulation of glycogen he production of bacteriocins (antimicrobial compounds),adherence
to the vaginal epithelia(competitive exclusion of other bacteria), and ability to competitively use
available nutrients (
by either Lactobacillus crispatus (CST I), L. gasseri (CST II), L. iners (CST III), or L. jensenii (CST V). Additionally, CST IV does not contain a
significant species or quantity of Lactobacillus but instead comprised of a polymicrobial mixture of strict and facultative anaerobes including
species Of the genera Gardnerella, Atopobium, Mobiluncus,Prevotella

Cervico-vaginal Microbiota in Infertility

BV-associated bacteria in the vagina have been shown to be associated with a reduced pregnancy rate

large cohort study of 266 healthy women initiating contraception and aged 18–35 years in Harare,

Zimbabwe, used quantitative polymerase chain reaction (PCR) measurement of vaginal bacteri

3)Insight into the ecology of vaginal bacterial through integrative analyses of metagenomic
and metatranscriptomic data
abundance of a species is not always indicative of its transcriptional activity and that impending changes in community composition can be
predicted from metatranscriptomic data

The ratio of a species relative activity divided by its relative abundance was referred to as its “relative expression

represents the extent to which the species is over- or under-represented in the metatranscriptome, as compared to the metagenome

under-expressive. G. vaginalis, A. vaginae, Finegoldia magna, Mobiluncus mulieris, and Streptococcus anginosus were more likely to be under-
expressive

over-expressive Sneathia amnii S. sanguinegens

The composition of a metagenome represents the relative abundances of taxa, whereas the composition of the metatranscriptome represents the
relative activities of taxa

L. iners and G. vaginalis both demonstrate lower expression of their cholesterol-dependent cytolysins when co-resident with Lactobacillus spp.
And higher expression when co-resident with other facultative and obligate anaerobes

The pathogenic potential of these species may depend on the communities in which they reside . Women whose
microbiota are largely depleted of lactobacilli are at greater risk to symptomatic bacterial vaginosis (BV), sexually
transmitted diseases, and preterm birth [26–30]. I
Integration of metagenomic and metatranscriptomic data identify potential species interactions

results from our analysis of metatranscriptomic data could guide the development of innovative strategies to
modulate the composition and activity of the vaginal microbiota to restore and maintain an optimal protective
environment.
 4) Vaginal Microbiome: Rethinking Health and Disease
human vagina and the bacterial community balanced mutualistic association

The indigenous bacterial communities, on the other hand, play a protective role in preventing colonization of the
host by potentially pathogenic organisms, including those responsible for symptomatic bacterial vaginosis, yeast
infections, sexually transmitted infections (STIs), and urinary tract infection

microorganisms benefit the host by producing lactic acid as a fermentation product that lowers the vaginal pH to~
3.5–4.5 (12).

vaginal communities experience chronic and acute disturbances caused by human behaviors, such as the use of
antibiotics, hormonal contraceptives and other methods of birth control, sexual activity, vaginal lubricants,
douching, and so forth, in addition to many other intrinsic factors such as the innate and adaptive immune systems of
hosts (

majority of microbial species (>99%) resist cultivation in the laboratory (culture-independent approaches based on
the analysis of 16S rRNA gene sequences Followed by,amplification of the 16S rRNA genes using primers that
anneal to highly conserved regions of the gene, followed by sequencing and classification of the phylotypes present

L. crispatus, Prevotella bivia, and Atopobium vaginae demonstrated that these key vaginal bacteria appear toregulate
the epithelial innate immunity in a species-specific manner

large-scale cross-sectional study of 396 healthy asymptomatic women revealed that L. iners was detected in 83.5%
of the subjects and dominated 34.1% of the communities analyzed L. crispatus, L. gasseri, and L. jensenii were
present in 64.5, 42.9, and 48.2% of the subjects and dominated in 26.2, 6.3, and 5.3% of the samples, respectively

Lactobacillus Species’ Antimicrobial Substances Production

Lactobacillus spp. produce hydrogen peroxide in vitro under aerobic conditions, which could inhibit colonization of
potential pathogenic bacteria but. L iners, fail to produce hydrogen peroxide. high concentrations of hydrogen
peroxide are even more toxic to vaginal Lactobacillus than to BVAB (8

suggests that lactic acid, not hydrogen peroxide, is more likely to contribute to the protective role of vaginal
microbiota

Facultative Anaerobes Mobiluncus, and Finegoldia Atopobium, Corynebacterium, Anaerococcus, Peptoniphilus,

Prevotella, Gardnerella, Sneathia, Eggerthella, , among

In clinical settings, BV is commonly diagnosed on the basis of the clinical criteria described by Amsel et al.),
wherein three of the following four symptoms must be evident: (a) a homogenous, white,noninflammatory discharge
that smoothly coats the vaginal walls; (b) the presence of clue cells (squamous epithelial cells covered with adherent
bacteria) on microscopic examination; (c) a vaginal fluid pH over 4.5; and (d ) a fishy odor of vaginal discharge
before or after addition of 10% KOH (potassium hydroxide

The dynamics of vaginal bacterial communities during the menstrual cycle, and the dramatic changes associated
with transitions between the physiological stages of a woman’s life span, from the first week of life to puberty,
reproductive years, andmenopause, are a reflection of the interplay of the mutualistic relationship between the
vaginal microbiota and its human host

Different aspect:sexual hormone levels, features of host physiology, and composition and functional output of the
vaginal microbiota
5) The vaginal microbiota, host defence and reproductive physiology
The interaction between the human host and the vaginal microbiota is highly dynamic. Major changes in the
vaginal physiology and microbiota over a woman’s lifetime are largely shaped by transitional periods such as
puberty, menopause and pregnancy, while daily fluctuations in microbial composition observed through
culture-independent studies
non-Lactobacillus vaginal microbiota may trigger immune responses as well as degrade the host mucosa,
processes that ultimately increase susceptibility to infections and contribute to negative reproductive outcomes
such as infertility and preterm birth
Gardnerella vaginalis or Lactobacillus iners) contribute to mucosa degradation and susceptibility to other
infection
observed that some vaginal microbial communities transitioned in and out of CST-IV.
some cases, CST transitions were triggered by menstruation or sexual behaviours,
The authors also showed how there were significant increases in IL-1α, IL-1β and TNF-α longitudinally in
subjects that transition from a CST-I, to CST-III and to a CST-IV CST-IV can clinically manifest as aerobic
vaginitis (AV).. AV is mainly differentiated from BV by presence of the an inflammatory response predominately
associated with aerobes, such as group B Streptococcus, Staphylococcus aureus, Escherichia coli, and Enterococcus
A number of immune factors including IL-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, TNF-a, IFN-., chemokine C-C
motif ligand 5 (CCL5) and SLPI have been variably and inconsistently associated with BV (

Lactic acid directly inhibit Chlamydia trachomatis infection (Gong et al. 2014), and potentially both HSV-2 and
HIV

Vvaginalis produces vaginolysin, a cholesterol-dependent cytolysin that could contribute to BV symptomology by

forming pores in the vaginal epithelium with or without CD59

BV-like vaginal microbiota are significantly more prevalent in women with tubal infertility when compared with
women with other causes of infertility, but is not associated with decreased conception rates

The early childhood vaginal microbiota Comprise a variety of anaerobes, diphtheroids, coagulase-negative staphy
lococci, and E. coli,whereas postmenopausalwomen often experience a loss of Lactobacillus spp

The human vaginal ecosystem is a dynamic environment in which microbes can affect host physiology but also
where host physiology can affect the composition and function of the vaginal microbiota. Species of Lactobacillus
have been historically associated with vaginal health

Lactobacillus products, such as lactic acid and bacteriocin among others, against mucus degradation and inhibition
of pathogens. The reported inconsistent innate immune response observed with non-Lactobacillus- or L. iners-
dominated micro biota (CST-IV, BV, AV and CST-III, respectively),

6)Interplay Between the Temporal Dynamics of the Vaginal Microbiota and

Human Papilloma virus Detection
temporal relationship between vaginal microbiota and human papilloma virus (HPV) detection.
CST was associated with changes in HPV status . Lactobacillus gasseri–dominated CSTs had the
fastest HPV remission rate, and a low Lactobacillus community with high proportions of the
genera Atopobium (CST IV-B) had the slowestn rate compared to L. crispatus–dominated CSTs.
L. gasseri was associated with increased clearance of detectable HPV.
persissistent detection of human papillomavirus (HPV) is associated with increased risk for
dysplasia and cervical malignancy
cross-sectional studies demonstrated a positive association between BV and cervical
HPV .participants who were HPV positive had a lower proportion of Lactobacillus spp. Than
HPV-negative women. In addition, the authors identified Fusobacteria, including Sneathia spp.
as key bacteria associated with HPV detection.
16S rRNA gene sequence analysis clustered into 6 CSTs: 4 dominated by either L. crispatus
(termed CST I), L. gasseri (CST II), L. iners (CST III), or L. jensenni (CST V), and 2 that lacked
substantial numbers of Lactobacillus
spp. (CST IV-A and IV-B). CST IV-A comprised a variety of anaerobic bacteria (including
Streptococcus and Prevotella),
whereas CST IV-B harbored higher proportions of the genera Atopobium, Gardnerella, and
Prevotella among others.
CST II (L. gasseri–dominated) was associated with the fastest HPV remission rate (HPV positive
to no detection) and CST
IV-B was associated with the slowest HPV remission rate copared to the CST
The vaginal microbiome may play a significant role in the control of latent HPV infection,

defense against incident HPV ac quisition,or increased clearance of HPV,

7)Association of HPV infection and clearance with cervicovaginal immunology

and the vaginal microbiota
HPV infection increase HIV risk. Repeat .HPVtesting was performed after 6 months. Cytobrush-
derived T cell and dendritic cell subsets were assessed by multiparameter flow cytometry. Undiluted
cervico-vaginal secretions to determine cytokine levels by multiplex ELISA, assess bacterial community
composition and structure by 16S rRNA gene sequence analysis. The majority of HPV infections are cleared
quickly by the host,2 but high-risk subtypes (HR-HPV) can persist..
inflammatory response may render individuals more susceptible to HIV infection
activated CD4 T CCR5,8 cells express elevated levels of the HIV co-receptor making them preferential HIV targets
endocervical CD4 T cell number did not differ

CD4þ between HPV negative participants (mean 1153 T cells/ cytobrush) and HPV positive participants (889
cells/cytobrush

participants who cleared HPV had a higher absolute number of endocervical Langerhans cells (5018 cells per
cytobrush) than both HPV negative women (635 cells per cytobrush, 0.015) and those with persistent HPV infection
(180 cells per cytobrush,

more participants with HPV infection had a cervico-vaginal microbiome consistent with CST-IV, com pared
with HPV-negative women (58.8% vs. 29.4%; 0.043

HPV infection was associated with alterations in the composition and structure of the vaginal microbiota

8)Perspective on human papillomavirus infection treatment by vaginal

microbiota
Almost all cervical cancers are caused by infection of high‐risk human papillomaviruses (hr‐
HPV), persistent genital high-risk HPV (hr-HPV) infection leads to about 99.7% of cervical
cancer cases. while HPV-16 and -18 are the most common cancer-causing types
HPV viral proteins
E6 and E7 are proved to be necessary for the carcinogenicity of cervical cancer cells
Transient and persistent HPV16 infections are found to be associated with a
vaginal microbiota dominated by non-Lactobacillus species. the Lactobacillus-dominated
microbiota results in a fast HPV clearance rate, while some anaerobic
bacteria such as Gardnerella vaginalis (G. vaginalis)restrain HPV clearance and promote the
development of CIN. cervical intraepithelial neoplasia
HPV E7 oncoprotein greatly inhibits some host defense peptide expression through NF-.B and
Wnt/ß-11 catenin signaling pathways.11 Consequently, amino acids that maintain the growth of
Lactobacillus species are remarkably down-regulated.
L. iners‐dominated microbiota
was more likely to be associated with vaginal dysbiosis
probiotic drink containing L. casei Shirota every day, results showed that 29% of patients in the
intervention
group cleared HPV,

9)Vaginal Microbiome Dysbiosis is Associated with the Different

Cervical Disease Status

four groups (NV + normal tissue with HPV infection , LSIL low-grade squamous intraepithelial
lesion , HSIL (high-grade squamous intraepithelial lesion), and CC). C The relative percentage of
HPV16, HPV18
and other hrHPV(High Risk)
characterized the vaginal microbiome of women with different status of cervical diseases, including (normal tissue
with HPV infection), low-grade squamous intraepithelial lesion (LSIL, high-grade squamous intraepithelial lesion
( found that higher diversity of microbiome with gradual depletion of Lactobacillus, especially L. crispatus, was
associated with the severity of cervical disease.. complete loss of
L. crispatus, was observed in women with CC… Cervical cancer (CC) is the fourth most common
Persistent high-risk human papillomavirus major risk factors
the enrichment of Sneathia, Megasphaera,Prevo tella, Gardnerella and/or bacterial vaginosis-
associated bacteria (BVAB) 1 and 2 were often associated with HPV infection
Colpos Copy and biopsy were carried out to confirm the cervical diseases status for women with
abnormal cytology and HPV-positive women with normal cytology
HPV16 was more prevalent in both the HSIL(high-grade squamous intraepithelial lesion) and CC group
Increased negative interactions between Lactobacillus and other bacteria
were observed in the HSIL group.CC group, the bacteria-bacteria interaction further developed
to a highly connected complex network with more non- Lactobacillus bacterias but no
Lactobacillus involved
Suggested that increased dysbiosis of vaginal microbiome was associated with high-grade
cervical lesions.
Lactobacillus iners is not only unable to produce D-lactic acid and H2O2, also generates
inerolysin, forms pores in vaginal epithelium to help virus infections
HPV16 infection was highly associated with higher vaginal microbiome diversity and the
depletion of Lactobacillus
Meantime, HPV16 positive rate was significantly
higher in the HSIL and CC groups than the Normal-LSIL
group, which supported that HPV16 as highest risk factor
The Vaginal Microbiome: II. Vaginal
Dysbiotic Conditions
The Vaginal Microbiome: III. The Vaginal Microbiome in Various
Urogenital Disorders
Candidiasis (vulvovaginal)
Vaginal Microbiota Changes Caused by HPV Infection in Chinese
Women
FIGURE 1. Depiction of a model for BV establishment and immunologic/physiological changes related to host-microbe
interactions. (a) In a healthy state, the vaginal microbiome is dominated by Lactobacillus spp. Lactobacilli produce lactic acid,
which lowers the vaginal pH and protects against invading pathogens and pathobionts. Epithelial cells produce low levels of
antimicrobial peptides (e.g. SLPI) and cytokines. Additionally, epithelial cells and immune cells contribute to homeostasis by
producing anti-inflammatory cytokines (e.g. IL-1RA). (b) Vaginal dysbiosis begins with initial colonization with G. vaginalis,
usually following a sexual exposure. G. vaginalis colonizes the vaginal epithelial cells, replaces lactobacilli, and provides
scaffolds for biofilm formation. Following G. vaginalis colonization, P. bivia is recruited to the biofilm and G. vaginalis and P.
bivia support each other’s growth. Both of these bacteria are capable of producing enzymes (i.e., sialidase) which may contribute
to mucus degradation and barrier disruption. No overt inflammation is observed, suggesting that they are able to evade the host
immune response. (c) Other secondary colonizers including A. vaginae and Sneathia spp. are recruited to the biofilm and
exfoliation of epithelial cells coated with polymicrobial biofilm occurs. Production of biogenic amines and other metabolites
contribute to elevated vaginal pH and BV symptoms. Epithelial cells and recruited immune cells produce proinflammatory
cytokines and chemokines.
Types and Causes of Vaginitis

Doctors refer to the various conditions that cause an infection or inflammation of the vagina as
"vaginitis." The most common kinds are:

 Bacterial vaginosis, inflammation of the vagina due to an overgrowth of bacteria. It typically

causes a strong fishy odor.
 Candida or "yeast" infection, an overgrowth of the fungus candida, which is normally found in
small amounts in the vagina.
 Chlamydia is the most common sexually transmitted infection (STI) in women, usually in those
ages 18 to 35 who have multiple sex partners.
 Gonorrhea is another common infection spread through sex. It often comes along with
chlamydia.
 Trichomoniasis is an infection spread by sex that’s caused by a parasite. It raises your risk for
other STIs.
 Viral vaginitis is inflammation caused by a virus, like the herpes simplex virus (HSV) or human
papillomavirus (HPV), which spread through sex. Sores or warts on the genitals can be painful.
increased risk of preterm birth, pelvic inflammatory disease, endometritis, cervical intraepithelial
neoplasia (CIN), and acquisition of STIs including HIV and human papillomavirus (HPV).

he Nugent score is based on gram-stained vaginal smear, with high numbers of lactobacilli species being
indicative of health, and their depletion coupled with increased numbers of small and/or curved gram
variable rods being indicative of BV.18

however, G. vaginalis was present in 98% of women with BV, A. vaginae in 92%, L. iners in 86%, and
Eggerthella species in 85%

Desquamative Inflammatory Vaginitis and Aerobic Vaginitis

Most often, isolated bacteria from patients with DIV have been Streptococcus agalactiae, Escherichia
coli, and Staphylococcus aureus.

he bacteria most often isolated in AV are Streptococcus species, S. aureus, S. epidermidis, S. anginosus,
E. coli, and E. faecali