Clinical and Personal Lubricants Impact the Growth of Vaginal Lactobacillus Species and Colonization of Vaginal Epithelial Cells: An in Vitro Study

Abstract

Background: Vaginal lubricants are commonly used during gynecological examinations, during sexual activities, or to alleviate vaginal dryness. Many lubricants contain potentially bacteriostatic or bactericidal agents (parabens, chlorhexidine gluconate, nonoxynol-9). Our objective was to evaluate the impact of lubricants that vary in formulation on the growth and viability of vaginal Lactobacillus species and vaginal epithelial cell (VEC) colonization in an in vitro model.

Methods: Growth curve, disk diffusion, and minimal inhibitory assays were used to determine the impact of lubricants or excipients on the growth of Lactobacillus crispatus, Lactobacillus gasseri, Lactobacillus jensenii, and Lactobacillus iners. L. crispatus strain was used in VEC colonization assays. Statistical differences were determined by analysis of variance.

Results: Lubricants containing chlorhexidine gluconate or nonoxynol-9 (N-9; Conceptrol, K-Y Jelly, and Surgilube) significantly inhibited Lactobacillus species growth (P < 0.05). In contrast, other clinical lubricants (E-Z Lubricating Jelly, McKesson Lubricating) and personal lubricants (Astroglide Liquid, Good Clean Love Almost Naked, K-Y Warming Jelly) did not exhibit this effect. Chlorhexidine gluconate had a detrimental effect on Lactobacillus growth and exhibited stronger antimicrobial activity compared with methylparaben and propylparaben (P < 0.0001). There were lubricants that did not induce cytotoxicity in VEC (Good Clean Love Almost Naked, E-Z Lubricating Jelly, McKesson Lubricating Jelly), but these products did substantially decrease the attachment of L. crispatus to VEC, particularly when VEC were preexposed to lubricants before inoculation with bacteria (P < 0.0001).

Conclusions: This in vitro model indicates that select vaginal lubricants, particularly those with chlorhexidine gluconate, have potentially adverse effects on women's health by reducing growth and recolonization of vaginal Lactobacillus species.

Figure 1.. Select lubricants (K-Y^™ Jelly, Surgilube® and Conceptrol®) inhibit the growth of vaginal Lactobacillus spp.

L. crispatus strain VPI 3199, L. crispatus strain JV-V01, L. gasseri strain JV-V03, L. jensenii strain JV-V16 and L. iners strain AB107 were grown in MRS or supplemented TSB broth with 10% (v/v) lubricants at 37°C under 5% CO₂ or anaerobic conditions. Bacterial growth was assessed by measuring optical densities at 600 nm (OD₆₀₀) at 4 h and 24 h post inoculation. The broth without any lubricant was used as a positive control. OD₆₀₀ measurements of each culture with lubricants were compared to cultures without lubricants at respective time points. Data are shown as means ± SE from at least three independent experiments. P values were calculated using one-way ANOVA with Dunnett post-test (* P<0.05; ** P<0.01; *** P<0.001; **** P<0.0001).

Figure 2.. Select lubricants (K-Y^™ Jelly, Surgilube® and Conceptrol®) exhibit bacteriostatic effect on vaginal Lactobacillus spp.

L. crispatus strain VPI 3199 (A), L. crispatus strain JV-V01 (B), L. gasseri strain JV-V03 (C), L. jensenii strain JV-V16 (D) and L. iners strain AB107 (E) were grown in MRS or supplemented TSB broth with 10% (v/v) lubricants at 37°C under 5% CO₂ or anaerobic conditions. Bacterial growth kinetics were assessed by measuring number of colony forming units (CFU) representing viable bacteria in the cultures in appropriate liquid media containing 10% solutions of lubricants at 4 and 24 h following the exposure to 10% (v/v) lubricants using standard plating assay. The broth without any lubricant was used as a positive control. Concentrations of viable bacteria in each culture were calculated as CFU/mL and compared to culture without lubricants at respective time points. Data are shown as means ± SE from three independent experiments. P values were calculated using one-way ANOVA with Dunnett post-test (* P<0.05; ** P<0.01; *** P<0.001; **** P<0.0001).

Figure 3.. Chlorhexidine gluconate exhibit stronger antimicrobial properties than parabens across all tested vaginal Lactobacillus spp.

A. The impact of each excipient on the growth of Lactobacillus spp. was tested using disk diffusion assay. Bacteria were grown on appropriate agar plates with 6-mm disks impregnated with 20% solutions (w/w) of methylparaben, propylparaben and chlorhexidine gluconate (CHG). Ethanol (solvent for parabens) and bleach was used as negative and positive controls, respectively. Zones of inhibition were recorded 24 h post inoculation and compared to a negative control. Data are shown as means ± SE from three independent experiments. P values were calculated using one-way ANOVA with Bonferroni post-test (** P<0.01; *** P<0.001; **** P<0.0001). B. Minimal inhibitory concentrations (MIC) of methylparaben, propylparaben and CHG were determined using the broth microdilution method. The MIC was defined as the lowest concentration of the excipient that inhibits the visible growth of the tested Lactobacillus spp.

Figure 4.. Lubricants reduce the colonization of vaginal epithelial cells (VEC) with L. crispatus particularly when VEC are colonized with bacteria following the lubricant exposure.

Three non-cytotoxic lubricants: Good Clean Love (GCL) Almost Naked, E-Z Lubricating Jelly, McKesson Lubricating Jelly) were tested to determine their impact on colonization of in vitro VEC model with L. crispatus strain JV-V01. A. VEC were pre-colonized with bacteria at the multiplicity of infection (MOI) 10 for 2 h prior to exposure to 10% (v/v) lubricants B. VEC were exposed to 10% (v/v) lubricants and immediately colonized with L. crispatus for 4 h or 24 h as described above. Colonization levels were reported as number of viable bacteria (CFU) attached to VEC per well. Data are shown as means ± SE from at least three independent experiments. P values were calculated using two-way ANOVA with Bonferroni post-test (** P<0.01; **** P<0.0001).