Vaginal flora

lactobacilli and a vaginal squamous cell.

The micro-organisms that colonize the vagina, collectively referred to as the vaginal microbiota or vaginal microflora, were discovered by the German gynecologist Albert Döderlein in 1892. The amount and type of bacteria present have significant implications for a woman's overall health. The primary colonizing bacteria of a healthy individual are of the genus lactobacillus,^[1] such as L. crispatus, and the lactic acid they produce (some species produce hydrogen peroxide or antibiotic), in combination with fluids secreted during sexual arousal, are greatly responsible for the characteristic scent associated with the vaginal area.

Vaginal microflora

Lactobacilli

The primary colonizing bacteria of a healthy individual are of the genus lactobacillus.^[1] Since the first description of lactobacilli by Döderlein, lactobacilli have been generally considered as the gatekeepers of the vaginal ecosystem. Lactobacilli have been shown to inhibit in vitro growth of pathogenic microorganisms, e.g. Bacteroides fragilis, Escherichia coli, Gardnerella vaginalis, Mobiluncus spp., Neisseria gonorrhoeae, Peptostreptococcus anaerobius, P. bivia and Staphylococcus aureus. It is generally accepted that this is achieved through the action of lactic acid primarily.^[2][3][4][5] Moreover, lactobacilli ensure long-term colonization of the vagina through adherence to vaginal epithelial cells, blocking the adherence of bacterial pathogens to the vaginal epithelium.^[6] Next to lactic acid production and competition for adherence, other antagonistic mechanisms consist of hydrogen peroxide (broad-spectrum antimicrobial) and bacteriocin (targetspecific antimicrobials) production.^{[7][8][9][10][11][12]}

The production of lactic acid

Low pH is generally accepted to be the main mechanism controlling the composition of the vaginal microflora. Although the lactic acid produced by lactobacilli contributes to the vaginal acidity, it is still not proven to be the primary source of low vaginal pH, but the fact remains that most lactobacilli thrive best at a pH < 4.5 .^[8][13][14]

Hydrogen peroxide

Production of H₂O₂ is a well-known mechanism for bacterial antagonism ^[15][16][17] , inhibiting growth of microorganisms via direct interaction or via human myeloperoxidase.^[18][19][20] Hydrogen peroxide-producing lactobacilli have been shown to inactivate HIV-1, herpes simplex virus type 2 (HSV-2), Trichomonas vaginalis, G. vaginalis, P. bivia and E. coli.^{[10][19][21][22][23][24][25][26]} Eschenbach et al.^[18] found that 96% of Lactobacillus species from a healthy vaginal ecosystem produced H₂O₂ (L. jensenii and L. vaginalis produce the highest levels of H₂O₂,^[9][27] whereas only 6% of the lactobacilli recovered from women with BV produced H₂O₂.^[18] In agreement with this, L. iners, most frequently associated with disturbed vaginal microflora,^[28][29] is a poor producer of H₂O₂.^[30][31] Vaginal colonization by H₂O₂-producing lactobacilli has been associated with a decrease in the occurrence of Bacterial vaginosis (BV).^[32] However, more recently O‘Hanlon et al.^[33] demonstrated that cervicovaginal fluid and semen have a significant H₂O₂-blocking activity and they later ^[10] demonstrated that physiological concentrations of H₂O₂ below 100 μM fail to inactivate any of the 17 tested BV-associated bacteria, e.g. A. vaginae, G. vaginalis, Mobiluncus spp., P. bivia, Prevotella corporis, Mycoplasma hominis, even in the presence of human myeloperoxidase, known to increase the microbicidal activity of H₂O₂.^[10] Only supraphysiologic concentrations of exogenous H₂O₂ (0.34% w/v, 100 mM) were sufficient to inactivate BV-associated bacteria at which concentration it more potently inactivated vaginal lactobacilli (L. crispatus, L. gasseri, L. iners and L. jensenii). A concentration of 100 mM H₂O₂ is approximately 50-fold higher than lactobacilli are capable of producing even under optimal aerobic, low-antioxidant conditions, and approximately 5,000-fold higher than the estimated H₂O₂ concentration in vivo. Even more remarkable, the addition of only 1% vaginal fluid blocked the microbicidal activity of 1 M H₂O₂. Possible explanations may be that cervicovaginal fluid and semen contain proteins, glycoproteins, polysaccharides, lipids, and other molecules with the potential to react with and inactivate H₂O₂. In addition, the vagina is hypoxic most of the time, whereas lactobacilli require oxygen to produce hydrogen peroxide. It is also remarkable that catalase, which provides bacteria protection against toxic H₂O₂, is absent in lactobacilli,^[18][21] and as such they would be unprotected against their own H₂O₂ production. In contrast, under optimal anaerobic growth conditions, physiological concentrations of lactic acid inactivated the BV-associated pathogens without affecting the vaginal lactobacilli.^[10][33] In summary, although the hydrogen peroxide production of lactobacilli has been considered as an important antimicrobial component, contributing to the colonization resistance provided by lactobacilli,^[9][12] and although there seems to be a link between H₂O₂-producing lactobacilli and normal vaginal microflora, recent data do not support this role for H₂O₂.^[10][33]

Bacteriocins

Vaginal lactobacilli produce several target specific antimicrobial peptides, i.e. bacteriocins such as lactocin 160 and crispasin A ^[8] ,^[34][35][36] with inhibitory activity ranging from narrow (closely related Lactobacillus species) ^[37] to broad (diverse groups of bacteria, including G. vaginalis and P. bivia),^{[4][36][38][39]} and bacteriocin-like substances, with a broader spectrum of activity than bacteriocins (e.g. a heat-resistant peptide produced by Lactobacillus salivarius subsp. salivarius CRL 1328).^[6][40] Several studies have indicated that the activity of bacteriocins is favored by low pH.^[4][41] In summary, the inhibitory substances produced by vaginal Lactobacillus species seem to be a primary factor in protecting the vaginal econiche, with organic acids, bacteriocins (and hydrogen peroxide) acting synergistically to prevent colonization by pathogenic anaerobes. Not all Lactobacillus spp. and not all strains within one Lactobacillus species exhibit all 3 mechanisms.^[8] It is of importance to note that only a select group of Lactobacillus species has been found to dominate the vaginal econiche of premenopausal women, i.e. L. crispatus, L. jensenii, L. iners, L. gasseri (and possibly L. vaginalis), as assessed through cultivation-dependent and cultivation-independent techniques.^{[28][29][30][42][43][44][45][46][47][48]} Vaginal lactobacilli have been shown to display a pronounced vaginotropism, and their pili act as ligands for attachment to glycolipid receptors of vaginal epithelial cells.^[49][50] The limited number of Lactobacillus spp. found in the human vagina is remarkable, which leads to the possibility that there are host factors that select for specific organisms, that these species have unusual characteristics that allow them to successfully colonize the vagina, or both .^[46] However, the vaginotropism, does not only apply to this selected group of lactobacilli that stand for a healthy vagina, but also for the bacterial species associated with BV.^[51] The microflora detected in the human genital and gut econiche do not appear to grow outside their host and probably are likely to rely on the close contact between parents and their children for transmission,^[51] e.g. mother to neonate transmission of genital microflora, most probably also with gut microflora homogenously distributed over the baby‘s body including skin, the oral cavity, nasopharynx, and feces.^[52] Molecular studies have shown that the intestinal microflora of identical and fraternal twins show greater similarities than between twins and their partners, this is probably due to a shared mother.^[50]

Other vaginal bacterial species

Several anaerobic bacterial species are frequently found in the vagina, such as the Gram positive cocci: Atopobium vaginae, Peptostreptococcus spp., Staphylococcus spp., Streptococcus spp., and Bacteroides spp., Fusobacterium spp., Gardnerella vaginalis, Mobiluncus, Prevotella spp., and Gram-negative enteric organisms, such as Escherichia coli.^{[28][29][44][53][54][55][56][57][58]} Also Candida albicans, Mycoplasma and Ureaplasma are frequently found in the vagina.^[49][53][57] Some of the obligate and facultative anaerobic bacteria are associated with BV.^[56] In the pyrosequencing study by Ravel et al.,^[44] comprising the vaginal microflora of 396 asymptomatic women, equally representing four ethnic groups (blacks, whites, Asian and Hispanics), five different communities of vaginal microflora were found, with four communities dominated by respectively Lactobacillus crispatus, L. gasseri, L. iners or L. jensenii and the fifth with higher proportions of strictly anaerobic bacteria, predominantly Megasphaera, Prevotella, Sneathia and Streptococcus spp. (designated the diverse group). Hummelen et al.^[43] pyrosequenced (16S rRNA gene) the vaginal microflora of 132 HIV positive Tanzanian women and they detected eight major clusters. The two clusters that were strongly associated with normal vaginal microflora, were dominated by L. iners respectively L. crispatus. Four clusters were strongly associated with BV, and were dominated by G. vaginalis, Lachnospiraceae, P. bivia or a mixture of different species. The remaining 2 clusters were represented by the combination L. iners/G. vaginalis and the combination Lachnospiraceae/Veillonellaceae. In addition, G. vaginalis and L. iners were detected in each sample, so they were proposed as core members of the vaginal microflora.^[43]

What is normal vaginal microflora?

Recent studies have provided more insight about the composition of vaginal microflora, and it is becoming clearer that a single core of normal vaginal microflora dominated by lactobacilli could be an oversimplification of the reality. More than 20 vaginal species of Lactobacillus have been detected, although only 4 to 5, namely L. crispatus, L. gasseri, L. iners, L. jensenii and possibly L. vaginalis dominate the vaginal econiche.^{[28][42][44][46][58][59][60][61]} Several studies have demonstrated that a significant proportion (7–33%) of healthy, asymptomatic women (especially black and Hispanic women ^[44][59] lack appreciable numbers of Lactobacillus species in the vagina,^{[43][44][46][58][59][62]} which may be replaced by other lactic acid-producing bacteria that contribute to vaginal acid production by fermentation, i.e. species from the genera Atopobium, Leptotrichia, Leuconostoc, Megasphaera, Pediococcus, Streptococcus and Weissella,^{[46][59][63][64]} and also Escherichia coli isolates have been shown to acidify their growth media in vitro through the production of lactate.^[65] C. albicans is also known to produce acid, to thrive in a low pH environment and to coexist with lactobacilli.^[66][67] Interestingly, all ethnic populations have vaginal microflora communities containing lactic acid producing bacteria ^[44][46] .^[64][68][69] This is indicative for the fact that, although the structure of the bacterial communities may differ between populations, health can be maintained provided the functions of these communities, for example the production of lactic acid, are maintained .^[46][70] This may be in agreement with the findings of previous studies ^[59][62] and calls for a reassessment of the predominant opinion that normal vaginal microflora is always characterized by the presence of high numbers of lactobacilli. Forney et al.^[71] postulated:” We suspect that the causes of and cures for BV will continue to be enigmatic until it is recognized that, although “normal and healthy” can be equated with high numbers of lactobacilli, the converse — “unhealthy” being equated with low numbers of or no lactobacilli — is not necessarily true”. This implies that not all communities may be equally resilient, so that if the resilience of a vaginal community is low then transitory changes in the structure of these communities may occur more readily in response to disturbances of various kinds, including menses, sexual intercourse, douching and contraceptive practices.^[59] Zhou et al.^[59] hypothesized: ―These differences in the structure and composition of microbial communities may underlie well-known differences in the susceptibility of women in these racial groups to BV and various vaginal infections‖.^[59][72][73] Although it can be agreed that we should be cautious about which vaginal microflora should be considered as disturbed, and although (lactic) acid is indeed produced by many species in each of these communities, the remark by Mirmonsef et al.^[74] that emphasis should be on vaginal pH and not on mere acid production in establishing normal vaginal microflora may be justified. The importance of maintaining low pH as a protective mechanism also follows from the fact that pH further decreases during pregnancy. Indeed, several studies suggest that BV without clinical symptoms is still a risk factor, not only for STD/HIV infection, but also for poor obstetric outcomes.^[66][75] The acid production may be counteracted differently by different types of vaginal microflora, whereby vaginal microflora not dominated by lactoabilli may produce more amines and ammonia, due to proteolytic activity, which is indeed a hallmark of disturbed vaginal microflora.

Menstruation

During menstruation, the concentration of vaginal microbiome is observed to decline.^[76] The effect of tampon use on vaginal flora is debated, but application of tampons appears not to significantly modify the balance of bacterial presence.

Disease prevention

A healthy vaginal microbiome aids in the prevention of bacterial vaginosis, yeast infections and other possible problems by maintaining an acidic pH (< 4.5) that is unfavourable for the growth of common pathogens, such as Gardnerella vaginalis. The lactobacilli present in a healthy vaginal microbiome also occupy the ecological niche that would otherwise be available for exploitation by pathogenic organisms. However, harmful bacteria or an imbalance in bacteria can lead to infection.

One method of reducing the risk of infection in the local area of the urethra is to urinate immediately after sex. Additionally, exclusive use of sterile contraceptives can assist in prevention of infection.

See also

• Bacterial vaginosis
• Human flora
• Human microbiome project
• Skin flora

References

External links

• Döderlein´s bacteria