Glycogen availability and pH variation in a medium simulating vaginal fluid influence the growth of vaginal Lactobacillus species and Gardnerella vaginalis

Abstract

Background Glycogen metabolism by Lactobacillus spp. that dominate the healthy vaginal microbiome contributes to a low vaginal pH (3.5–4.5). During bacterial vaginosis (BV), strict and facultative anaerobes including Gardnerella vaginalis become predominant, leading to an increase in the vaginal pH (> 4.5). BV enhances the risk of obstetrical complications, acquisition of sexually transmitted infections, and cervical cancer. Factors critical for the maintenance of the healthy vaginal microbiome or the transition to the BV microbiome are not well defined. Vaginal pH may affect glycogen metabolism by the vaginal microflora, thus influencing the shift in the vaginal microbiome. Results The medium simulating vaginal fluid (MSVF) supported growth of L. jensenii 62G, L. gasseri 63 AM, and L. crispatus JV-V01, and G. vaginalis JCP8151A at specific initial pH conditions for 30 d. L. jensenii at all three starting pH levels (pH 4.0, 4.5, and 5.0), G. vaginalis at pH 4.5 and 5.0, and L. gasseri at pH 5.0 exhibited the long-term stationary phase when grown in MSVF. L. gasseri at pH 4.5 and L. crispatus at pH 5.0 displayed an extended lag phase over 30 d suggesting inefficient glycogen metabolism. Glycogen was essential for the growth of L. jensenii, L. crispatus, and G. vaginalis; only L. gasseri was able to survive in MSVF without glycogen, and only at pH 5.0, where it used glucose. All four species were able to survive for 15 d in MSVF with half the glycogen content but only at specific starting pH levels – pH 4.5 and 5.0 for L. jensenii, L. gasseri, and G. vaginalis and pH 5.0 for L. crispatus. Conclusions These results suggest that variations in the vaginal pH critically influence the colonization of the vaginal tract by lactobacilli and G. vaginalis JCP8151A by affecting their ability to metabolize glycogen. Further, we found that L. jensenii 62G is capable of glycogen metabolism over a broader pH range (4.0–5.0) while L. crispatus JV-V01 glycogen utilization is pH sensitive (only functional at pH 5.0). Finally, our results showed that G. vaginalis JCP8151A can colonize the vaginal tract for an extended period as long as the pH remains at 4.5 or above.

Under conditions closely mimicking vaginal fluid, Lactobacillus jensenii strain 62B produces a bacteriocin-like inhibitory substance that targets and eliminates Gardnerella species

“…As MRSB does not support the growth of Gardnerella spp. [51], we resuspended the retentates recovered from 10 kDa MWCO columns (fLj-MRSB-CFS24 and fMRSB) in NYCB and added them to additional NYCB (1 : 6 v/v) inoculated with Gp JCP8151B. As expected, the fMRSB (retentate resuspended in NCYB) did not support further growth of Gp JCP8151B; at 24 hpi, the initial inoculum was reduced by one log but not eliminated ().…”

“…Since the greatest reduction in c.f.u. occurred in the Gp JCP8151B-Lj 62B co-culture (), and since we had previously found that the growth of Lj 62B was more robust in MSVF at pH 5.0 [51], we continued our analyses using Lj 62B. To investigate this phenomenon further, and to determine if this effect might occur at an earlier time point during the co-culturing, we repeated the experiment but obtained samples at 16, 20, 22, and 24 hpi.…”

“…As shown in , Lj 62B viability gradually increased between 16 and 24 hpi. Our recent growth curve analysis of Lj 62B in MSVF over 30 d showed that growth was exponential at 24 to 48 hpi followed by a prolonged stationary phase during which Lj 62B maintain viability without a significant increase in growth over the remaining 28 d [51]. As seen with BLIS produced by L.…”

“…gasseri 63 AM, and L. crispatus JV-V01 individually under different pH conditions for an extended period (30 d) [51]. In this study, we used the Transwell two-chamber co-culture technique, which prevents direct contact between the bacteria in the upper and lower chambers, to determine if extracellular products synthesized by these vaginal Lactobacillus spp.…”

“…in vivo , it is important to utilize an in vitro medium that closely mimics the environment within the vagina and supports the growth of both lactobacilli and Gardnerella spp. Such a medium, the medium simulating vaginal fluid (MSVF) [ 50 ], supported the viability of G. piotii (Gp) JCP8151B and the vaginal lactobacilli L. jensenii 62B (Lj 62B), L. gasseri 63 AM, and L. crispatus JV-V01 individually under different pH conditions for an extended period (30 d) [ 51 ]. In this study, we used the Transwell two-chamber co-culture technique, which prevents direct contact between the bacteria in the upper and lower chambers, to determine if extracellular products synthesized by these vaginal Lactobacillus spp.…”