the role of acidification in the inhibition of neisseria gonorrhoeae by vaginal lac to bacilli...

8/3/2019 The Role of Acidification in the Inhibition of Neisseria Gonorrhoeae by Vaginal Lac to Bacilli During Anaerobic Growth

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Journal Reading Annals Of Clinical Microbiology and Antimicrobials 2011, 10:8

By : Diana Astari

Moderator : Dewi Kartika T.

Day/Date : Wednesday/ October, 19th 2011

Clinical Pathology Department, Faculty Of Medicine Universitas Padjajaran/RSUP Dr.

Hasan Sadikin BandungThe Role Of Acidification In The Inhibition ofNeisseria gonorrhoeae By

Vaginal Lactobacilli During Anaerobic Growth

Michelle A Graver, Jeremy J Wade

BACKGROUND

The majority of healthy women harbor lactobacillli (LB) in the vagina at counts of 107-109

cfu/gm secretions, protect the female genital tract to produce lactic acid. LB may also inhibit

opportunist bacteria through production of bacteriocins, hydrogen peroxide (H2O2) or local

immune response.

In bacterial vaginosis (BV) is replacement of normal vagina LB by a mixed flora of

anaerobic bacteria and may increase susceptibility to infection with sexually-transmitted

pathogens includingNeisseria gonorrhoeae (NG), a facultative anaerobe.

Bacterial interference between LB and NG growing anaerobically in liquid medium has

not been studied. By co-cultivating LB and NG, this study sought to identify the relative

contribution of LB acidification and H2O2production to any inhibition of NG under anaerobic

conditions.

METHODS

Media

A defined medium rendered anaerobic with Oxyrase for Broth (Oxyrase Inc, Ohio,

USA) was used, with ad without MES buffer adjusted to an intial pH 6.5 with 1 M HCl.

Bacterial Strains

The strains studied included three LB : Lactobacillus crispatus (LC), Lactobacillusgasseri (LG), andLactobacillus jensenii (LJ) distinguishable by auxotyping by the method ofCopley & Egglestone. Fusobacterium nucleatum (FN) was included as an indicator of

anaerobiosis.

Preparation Of Inocula

Inocula were prepared from overnight growth on chocolate agar in 5% CO2 at 37C for

NG, from blood agar incubated anaerobically overnight at 37C for FN, and from blood agar

incubated in 5% CO2 at 37C for 48 h for LB (media from Oxoid, Basingstoke, UK).

Monomicrobial Culture


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Monomicrobial growth studies were done to ensure that all bacteria were capable of

growth in medium, to identify any effect of MES buffer on growth, and to confirm

anaerobiosis by growth of the anaerobe FN.

Co-cultivation

Co-cultivation experiments were designed to demostrate whether high concentrations of

LB affect the growth of NG and if the addition of a pH buffer had any influence.

Sampling, Colony Counts And Growth Curves

Capped bijoux were incubated at 37C, and at times 15 h, 20 h and 25 h were examined

and confirmed reduction of inoculation and up to at least 25 h. Colonies were counted using

an aCOLyte colony counter (Don Whitley, UK). To demonstrate growth in the media, and

any effect of MES buffer, growth curves were prepared for the FN, NG and LB with and

without MES, using the median bacterial count at each time point.

RESULTS

The median counts (median log10 cfu/mL) for FN at times 0 and 25 h were 5.27 and 8.2,

respectively, confirming anaerobiosis. All LB and NG grew in media with and without MES

buffer. LB at the high inoculum grew over the first 15 h after inoculation and remained viable

thereafter. The addition of MES had negligible effect on growth.

In co-cultivation experiments, the median log10 cfu/mL inocula for LB and NG were 6.78

and 4.0, respectively. In the absence of MES buffer both LC and LG reduced the pH by > 1

point by 25 h; MES resisted this acidification, with a pH fall of


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used in for bacterial interference studies. This inhibitory actions appears to be primarily due

to acidification rather than H2O2 production demostrated by the correlation between the

degree of inhibition and the acidification capabilities of individual LB, the effects of buffer

and lacks of H2O2production. This study remains possible that a bacteriocin produced at, or

activated by, a lower pH could account. These study suggest that those seeking to select LB

for probiotic use in BV should address the acidification potential of candidate LB

CONCLUSION

During anaerobic growth, inhibition of NG by the vaginal LB tested was primarily due to

acidification, as evidenced by reduced inhibition in the presence of buffer. There was no

evidence of a specific mechanism of inhibition other than acid production, H2O2 was not

produce under these conditions. The acidification potential of vaginal lactobacilli under

anaerobic conditions may be an important characteristic conferring protection against NG

infection.

The Role Of Acidification In The Inhibition ofNeisseria gonorrhoeae By Vaginal

Lactobacilli During Anaerobic Growth

Background:

- Vaginal lactobacilli (LB) protect the female genital tract by producing lactic acid.

- LB inhibit opportunist bacteria through production bacteriocins, H2O2, or local immune response.

- Bacterial vaginosis (BV) is replacement of normal vagina LB by a mixed flora of anaerobic

bacteria and may increase susceptibility to infection with sexually-transmitted pathogens including

Neisseria gonorrhoeae (NG)

By co-cultivating NG in the presence of LB, to identify the relative contributions ofacidificat2ion and H2O2production to any growth inhibition of NG.

Methods :


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Three strains of NG by auxotyping grown in the presence of high concentrations of three vaginal LB

(L. Crispatus, L.gasseri and L.jensenii) in anaerobic liquid medium, wit or without MES buffer.

Fusobacterium nucleatum as an indicator anaerobiosis.

Bacterial counts performed at 15, 20 and 25 h; at 25 h pH and H2O2 concentration

Result:

Growth ofF.nucleatum confirmed anaerobiosis

All bacteria grew in anaerobic liquid medium and addition MES buffer had effect on growth.

L.crispatus and L.gasseri produced acidification and reduction in growth of NG.

L.jensenii produced less acidification and did not inhibit NG.

H2O2 not detected.

Discussion:

LB, normal vaginal flora plays a key role in maintaining vaginal health.

BV characterized by loss of the protective LB flora in the vagina & problem as risk factor infections

gonorrhoeae.

H2O2production is an important protective characteristic of LB comprising in normal vaginal flora.

Antibacterial effect H2O2permit LB to suppress other bacteria and control the vaginal flora.

NG can growth in anaerobic conditions and inhibit by vaginal LB growing.

Inhibitory appears due to acidification rather than H2O2production.

Correlation between the degree inhibition and the acidification individual LB, the effects of buffer,

and lack of H2O2production.

Conclusion :

During anaerobic growth inhibition of NG by LB due to acidification and abrogated by the presence

buffer.

H2O2 not produced

The acidification potential of vaginal lactobacilli under anaerobic conditions may be an

important characteristic conferring protection against NG infection.

the role of acidification in the inhibition of neisseria gonorrhoeae by vaginal lac to bacilli...

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