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The role of acidification in the inhibition of Neisseria gonorrhoeae by vaginal lactobacilli during anaerobic growth.

Graver MA, Wade JJ - Ann. Clin. Microbiol. Antimicrob. (2011)

Bottom Line: Fusobacterium nucleatum was used as an indicator of anaerobiosis.All bacteria grew in the anaerobic liquid medium and the addition of MES buffer had negligible effect on growth.There was no evidence of a specific mechanism of inhibition other than acid production under these conditions and, in particular, hydrogen peroxide was not produced.

View Article: PubMed Central - HTML - PubMed

Affiliation: Medical Microbiology, King's College Hospital, Denmark Hill, London, UK.

ABSTRACT

Background: Vaginal lactobacilli protect the female genital tract by producing lactic acid, bacteriocins, hydrogen peroxide or a local immune response. In bacterial vaginosis, normal lactobacilli are replaced by an anaerobic flora and this may increase susceptibility to Neisseria gonorrhoeae, a facultative anaerobe. Bacterial interference between vaginal lactobacilli and N. gonorrhoeae has not been studied in liquid medium under anaerobic conditions. By co-cultivating N. gonorrhoeae in the presence of lactobacilli we sought to identify the relative contributions of acidification and hydrogen peroxide production to any growth inhibition of N. gonorrhoeae.

Methods: Three strains of N. gonorrhoeae distinguishable by auxotyping were grown in the presence of high concentrations (107-108 cfu/mL) of three vaginal lactobacilli (L. crispatus, L. gasseri and L. jensenii) in an anerobic liquid medium with and without 2-(N-morpholino)-ethanesulfonic (MES) buffer. Fusobacterium nucleatum was used as an indicator of anaerobiosis. Bacterial counts were performed at 15, 20 and 25 h; at 25 h pH and hydrogen peroxide concentrations were measured.

Results: Growth of F. nucleatum to >108 cfu/mL at 25 h confirmed anaerobiosis. All bacteria grew in the anaerobic liquid medium and the addition of MES buffer had negligible effect on growth. L. crispatus and L. gasseri produced significant acidification and a corresponding reduction in growth of N. gonorrhoeae. This inhibition was abrogated by the addition of MES. L. jensenii produced less acidification and did not inhibit N. gonorrhoeae. Hydrogen peroxide was not detected in any experiment.

Conclusions: During anaerobic growth, inhibition of N. gonorrhoeae by the vaginal lactobacilli tested was primarily due to acidification and abrogated by the presence of a buffer. There was no evidence of a specific mechanism of inhibition other than acid production under these conditions and, in particular, hydrogen peroxide was not produced. The acidification potential of vaginal lactobacilli under anaerobic conditions may be their most important characteristic conferring protection against N. gonorrhoeae infection.

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Growth curves for N. gonorrhoeae. Growth of N. gonorrhoeae alone (black lines; as in Figure 1B) and in the presence of high inoculum of lactobacilli: L. crispatus (red lines; A and C), L. gasseri (blue lines; B and E), or L. jensenii (green lines; C and F), with (A-C) and without (D-F) MES buffer. Lines connect median value for triplicate experiments. Numbers in boxes above X axis show change in pH units of medium containing lactobacilli between times 0 and 25 h.
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Figure 2: Growth curves for N. gonorrhoeae. Growth of N. gonorrhoeae alone (black lines; as in Figure 1B) and in the presence of high inoculum of lactobacilli: L. crispatus (red lines; A and C), L. gasseri (blue lines; B and E), or L. jensenii (green lines; C and F), with (A-C) and without (D-F) MES buffer. Lines connect median value for triplicate experiments. Numbers in boxes above X axis show change in pH units of medium containing lactobacilli between times 0 and 25 h.

Mentions: In co-cultivation experiments, the median log10cfu/mL inocula for LB and NG were 6.78 and 4.0, respectively (Figure 1D & Figure 2). In the absence of MES buffer, both LC and LG reduced the pH by >1 pH point by 25 h; MES resisted this acidification, with a pH fall of <0.5 pH value for both. In contrast, LJ reduced the pH by only 0.7 pH point with a correspondingly lower pH fall in the presence of MES. Although the three NG grew in the presence of a high concentration of each LB, NG growth with LC and LG was reduced compared to growth without LB, and MES abrogated this inhibition. The presence of LJ had no effect on NG whether MES was included or not. H2O2 was not detected at 25 h in any experiment.


The role of acidification in the inhibition of Neisseria gonorrhoeae by vaginal lactobacilli during anaerobic growth.

Graver MA, Wade JJ - Ann. Clin. Microbiol. Antimicrob. (2011)

Growth curves for N. gonorrhoeae. Growth of N. gonorrhoeae alone (black lines; as in Figure 1B) and in the presence of high inoculum of lactobacilli: L. crispatus (red lines; A and C), L. gasseri (blue lines; B and E), or L. jensenii (green lines; C and F), with (A-C) and without (D-F) MES buffer. Lines connect median value for triplicate experiments. Numbers in boxes above X axis show change in pH units of medium containing lactobacilli between times 0 and 25 h.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3045876&req=5

Figure 2: Growth curves for N. gonorrhoeae. Growth of N. gonorrhoeae alone (black lines; as in Figure 1B) and in the presence of high inoculum of lactobacilli: L. crispatus (red lines; A and C), L. gasseri (blue lines; B and E), or L. jensenii (green lines; C and F), with (A-C) and without (D-F) MES buffer. Lines connect median value for triplicate experiments. Numbers in boxes above X axis show change in pH units of medium containing lactobacilli between times 0 and 25 h.
Mentions: In co-cultivation experiments, the median log10cfu/mL inocula for LB and NG were 6.78 and 4.0, respectively (Figure 1D & Figure 2). In the absence of MES buffer, both LC and LG reduced the pH by >1 pH point by 25 h; MES resisted this acidification, with a pH fall of <0.5 pH value for both. In contrast, LJ reduced the pH by only 0.7 pH point with a correspondingly lower pH fall in the presence of MES. Although the three NG grew in the presence of a high concentration of each LB, NG growth with LC and LG was reduced compared to growth without LB, and MES abrogated this inhibition. The presence of LJ had no effect on NG whether MES was included or not. H2O2 was not detected at 25 h in any experiment.

Bottom Line: Fusobacterium nucleatum was used as an indicator of anaerobiosis.All bacteria grew in the anaerobic liquid medium and the addition of MES buffer had negligible effect on growth.There was no evidence of a specific mechanism of inhibition other than acid production under these conditions and, in particular, hydrogen peroxide was not produced.

View Article: PubMed Central - HTML - PubMed

Affiliation: Medical Microbiology, King's College Hospital, Denmark Hill, London, UK.

ABSTRACT

Background: Vaginal lactobacilli protect the female genital tract by producing lactic acid, bacteriocins, hydrogen peroxide or a local immune response. In bacterial vaginosis, normal lactobacilli are replaced by an anaerobic flora and this may increase susceptibility to Neisseria gonorrhoeae, a facultative anaerobe. Bacterial interference between vaginal lactobacilli and N. gonorrhoeae has not been studied in liquid medium under anaerobic conditions. By co-cultivating N. gonorrhoeae in the presence of lactobacilli we sought to identify the relative contributions of acidification and hydrogen peroxide production to any growth inhibition of N. gonorrhoeae.

Methods: Three strains of N. gonorrhoeae distinguishable by auxotyping were grown in the presence of high concentrations (107-108 cfu/mL) of three vaginal lactobacilli (L. crispatus, L. gasseri and L. jensenii) in an anerobic liquid medium with and without 2-(N-morpholino)-ethanesulfonic (MES) buffer. Fusobacterium nucleatum was used as an indicator of anaerobiosis. Bacterial counts were performed at 15, 20 and 25 h; at 25 h pH and hydrogen peroxide concentrations were measured.

Results: Growth of F. nucleatum to >108 cfu/mL at 25 h confirmed anaerobiosis. All bacteria grew in the anaerobic liquid medium and the addition of MES buffer had negligible effect on growth. L. crispatus and L. gasseri produced significant acidification and a corresponding reduction in growth of N. gonorrhoeae. This inhibition was abrogated by the addition of MES. L. jensenii produced less acidification and did not inhibit N. gonorrhoeae. Hydrogen peroxide was not detected in any experiment.

Conclusions: During anaerobic growth, inhibition of N. gonorrhoeae by the vaginal lactobacilli tested was primarily due to acidification and abrogated by the presence of a buffer. There was no evidence of a specific mechanism of inhibition other than acid production under these conditions and, in particular, hydrogen peroxide was not produced. The acidification potential of vaginal lactobacilli under anaerobic conditions may be their most important characteristic conferring protection against N. gonorrhoeae infection.

Show MeSH
Related in: MedlinePlus