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Sihn and Yang: The Growth Inhibition Effect on the Causative Bacteria of Bacterial Vaginosis by Bacterial Strains Isolated from the Vagina of a Healthy Woman
Original Article

J Bacteriol Virol 2014;44(3):244-251.

Published online: 9 February 2014

DOI: https://doi.org/10.4167/jbv.2014.44.3.244

The Growth Inhibition Effect on the Causative Bacteria of Bacterial Vaginosis by Bacterial Strains Isolated from the Vagina of a Healthy Woman

Wan-Jin Sihn1, Nam-Woong Yang2, *

1Department of Alternative Complementary Medicine, Graduate School Chosun University, Gwangju, Korea

2Department of Microbiology, College of Medicine, Chosun University, Gwangju, Korea

*Corresponding author: Nam-Woong Yang. Department of Microbiology, College of Medicine, Chosun University, 309 Pilmun-daero Dong-gu Gwangju 501-759, Korea. Phone: +82-62-230-6350, Fax: +82-62-232-3125, e-mail: nwyang@chosun.ac.kr

** This study was supported by research fund from Chosun University, 2013.

Received 15 May 201422 July 2014       Accepted 24 July 2014

Copyright © 2014 Journal of Bacteriology and Virology

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Two Gram-positive rod strains isolated from the healthy vagina of a woman were tested for the possibility as probiotics. One strain was identified as Steroidobacter denitrificans (YH1) and the other as Lactobacillus crispatus (YH2) by 16S rRNA partial sequencing. The Casman agar and Man-Rogosa-Sharpe (MRS) agar were mixed in same quantity, supplemented with 5% human rbc lysate (CMB agar). The Wilkins-Chalgren agar and MRS agar were mixed in same quantity (WCM agar). Gardnerella vaginalis was cultured in Casman broth, supplemented with 5% human rbc lysate and 1,000 x-diluted with normal saline. Bacteroides fragilis, Mobiluncus mulieris and Peptostreptococcus asaccharolyticus were cultured in Wilkins-Chalgren anaerobe broth and 2,000x-diluted. S. denitrificans YH1 and L. crispatus YH2 were cultured in MRS broth anaerobically and 100x-diluted. The diluted suspensions of B. fragilis, M. mulieris and P. asaccharolyticus were inoculated on WCM agar and G. vaginalis on CMB agar by cotton swabs. Ten μl aliquots of YH1 and YH2 were inoculated on the center of WCM agar and CMB agar. The growth inhibition zone diameters of B. fragilis, G. vaginalis, M. mulieris and P. asaccharolyticus by YH1 were 35 mm, 35 mm, 25 mm and 60 mm. The inhibition diameters by YH2 were 25 mm, 30 mm, 20 mm and 40 mm, respectively. These results implicate that S. denitrificans YH1 can be the stronger probiotics for the treatment of bacterial vaginosis than L. crispatus, compared inhibition zone diameters by YH1 and YH2.

Keywords: Bacterial vaginosis, Lactobacillus crispatus, Probiotics, Steroidobacter denitrificans

REFERENCES

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Figure 1.
Sample A shows the 99% similarity to Steroidobacter denitrificans accession No. EF605262 in the unrooted neighbour-joining phylogenetic tree based on 16s rRNA gene sequence comparison.
jbv-44-244f1.tif
Figure 2.
Sample B shows the 99% similarity to Lactobacillus crispatus accession No. AF257097 in the unrooted neighbour-joining phylogenetic tree based on 16s rRNA gene sequence comparison.
jbv-44-244f2.tif
Figure 3.
Growth inhibition zones by S. denitrificans YH1. A, B, C, and D is B. fragilis, G. vaginalis, M. mulieris, and P. asaccharolyticus in regular order.
jbv-44-244f3.tif
Figure 4.
Growth inhibition zones by L. crispatus YH2. A, B, C, and D is B. fragilis, G. vaginalis, M. mulieris, and P. asaccharolyticus in regular order.
jbv-44-244f4.tif
Table 1.
Carbohydrate biochemical test results of L. crispatus YH2 by API CHL kit
No Lactobacillus crispatus YH2
 No Lactobacillus crispatus YH2
API CHL API CHL
0 Control − 25 Esculine +
1 Glycerol − 26 Salicine −
2 Erythritol − 27 Cellobiose −
3 D-Arabinose − 28 Maltose +
4 L-Arabinose − 29 Lactose +
5 Ribose − 30 Melibiose −
6 D-Xylose − 31 Saccharose +
7 L-Xylose − 32 Trehalose −
8 Adonitol − 33 Inuline −
9 β-Methyl-xyloside − 34 Melezitose −
10 Galactose + 35 D-Raffinose −
11 D-Glucose + 36 Amidon −
12 D-Fructose + 37 Glycogen +
13 D-Mannose + 38 Xylitol −
14 L-sorbose − 39 β-Gentiobiose −
15 Rhamnose − 40 D-Turanose −
16 Dulcitol − 41 D-Lyxose −
17 Inositol − 42 D-Tagatose −
18 Mannitol − 43 D-Fucose −
19 Sorbitol − 44 L-Fucose −
20 α-ethyl-D-mannoside − 45 D-Arabitol −
21 α-Methyl-D-glucoside − 46 L-Arabitol −
22 N-acetyl glucosamine + 47 Gluconate −
23 Amygdaline − 48 2-Keto-gluconate −
24 Arbutine − 49 5-Keto-gluconate −
Table 2.
Carbohydrate biochemical test results of S. denitrificans YH1 by API CHL kit
No Steroidobacter denitrificans YH1
 No Steroidobacter denitrificans YH1
API CHL API CHL
0 Control − 25 Esculine +
1 Glycerol − 26 Salicine +
2 Erythritol − 27 Cellobiose +
3 D-Arabinose − 28 Maltose +
4 L-Arabinose − 29 Lactose +
5 Ribose − 30 Melibiose −
6 D-Xylose − 31 Saccharose +
7 L-Xylose − 32 Trehalose +
8 Adonitol − 33 Inuline −
9 β-Methyl-xyloside − 34 Melezitose −
10 Galactose + 35 D-Raffinose −
11 D-Glucose + 36 Amidon +
12 D-Fructose + 37 Glycogen +
13 D-Mannose + 38 Xylitol −
14 L-sorbose − 39 β-Gentiobiose −
15 Rhamnose − 40 D-Turanose −
16 Dulcitol − 41 D-Lyxose −
17 Inositol − 42 D-Tagatose −
18 Mannitol + 43 D-Fucose −
19 Sorbitol − 44 L-Fucose −
20 α-ethyl-D-mannoside − 45 D-Arabitol −
21 α-Methyl-D-glucoside − 46 L-Arabitol −
22 N-acetyl glucosamine + 47 Gluconate −
23 Amygdaline − 48 2-Keto-gluconate −
24 Arbutine − 49 5-Keto-gluconate −
Table 3.
Enzyme activity results of Steroidobacter denitrificans YH1 by API ZYM kit
No Analysis items Results
0 Control −
1 Alkaline phosphatase +
2 Esterase (C4) −
3 Esterase Lipase (C8) −
4 Lipase (C14) −
5 Leucine arylamidase −
6 Valine arylamidase −
7 Crystine arylamidase −
8 Trypsin +
9 α-chymotrypsin −
10 Acid phosphatase −
11 Naphthol-AS-B1-phosphohydrolase −
12 α-galactosidase −
13 β-galactosidase +
14 β-glucuronidase −
15 α-glucosidase +
16 β-glucosidase +
17 N-acetyl-β-glucosaminidase −
18 α-mannosidase −
19 α-fucosidase −
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