• Journal of Dental Rehabilitation and Applied Science
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• v.37 no.4
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• pp.244-250
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• 2021

Purpose: The purpose of this study was to examine effects of culture conditions on the growth and antibacterial activity of Streptococcus salivarius K12. Materials and Methods: S. salivarius K12 was cultivated in medium containing animal and plant protein or in medium of neutral and acidic conditions. The growth of S. salivarius K12 was measured every 2 hours by a spectrophotometer. The antimicrobial activity of S. salivarius K12 against Streptococcus mutans and Porphyromonas gingivalis was investigated by the susceptibility assay using the spent culture medium. Results: the growth of S. salivarius K12 showed faster in medium containing plant protein and neutral pH condition. The antimicrobial and antifungal activity of S. salivarius K12 appeared stronger in medium containing plant protein than animal proteins. Conclusion: For application of S. salivarius K12 to bacterial oral disease, co-substances may be needed for S. salivarius K12 to colonize in the oral cavity and enhance the antimicrobial activity.

• Journal of the Korean Society of Food Science and Nutrition
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• v.38 no.12
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• pp.1811-1814
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• 2009

The effect of fermentation with mixed cultures of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus salivarius subsp. thermophilus on the microbiological characteristics of oat extract was investigated. Changes in pH, titratable acidity and viable cell populations indicated that growth was better in mixed cultures of L. delbrueckii subsp. bulgaricus and S. salivarius subsp. thermophilus. Growth of S. salivarius subsp. thermophilus in oat extract was more rapid than growth of L. delbrueckii subsp. bulgaricus. Cooperative interaction between two cultures during fermentation of oat extract as in yogurt from cow's milk was observed, but the intensity was relatively weak.

• Food Science of Animal Resources
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• v.31 no.4
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• pp.609-616
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• 2011

In this study, a LCH1227 bacterial strain that possesses anti-listerial activity was isolated from fermented food and identified as Lactobacillus salivarius LCH1227 based on its morphological and biochemical properties, as well as its 16S rRNA gene sequences. Anti-listerial substance also inhibited the growth of various Gram-positive bacteria, such as vancomycinresistant Enterococcus faecalis, Streptococcus agalactiae, Bacillus cereus, Lactobacillus fermentum. The highest level of production of antimicrobial substances from L. salivarius LCH1227 occurred during the early stationary phase. The antilisterial activity was found to be stable over a broad range of pH values (2.0-12.0) and after heat treatment. However, it was inactivated by proteolytic enzymes, indicating its proteinaceous nature. The apparent molecular mass of the partially purified anti-listerial substance, as measured by Tricine-SDS-PAGE, was approximately 5 kDa.

• Journal of the korean academy of Pediatric Dentistry
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• v.27 no.1
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• pp.15-23
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• 2000

Streptococcus salivarius is a normal inhabitant in the human oral cavity. Streptococcus salivarius 119 in this study was isolated from the oral cavity of child and identified, and its action mechanism on the formation of denal plaque by Streptococcus matans was studied. 1. The optical density of absorbance at 550 nm was 0.327 in the culture of Streptococcus mutans in disposable cuvette, whereas being 0.119 in the combined culture of Streptococcus mutans and Streptococcus salivarius 119. 2. The mean weight of produced artificial plaque on the wires in the beaker was 84.7mg in culture of Streptococcus mutans only, whereas being reduced to 12.3mg in the combined culture of Streptococcus mutans and Streptococcus salivarius 119. 3. When Streptococcus mutans was cultured in the media containing culture supernatant of Streptococcus salivarius 119 in BHI broth, the mean weight of produced artificial plaque was 100.5mg on the wires, whereas being reduced to 20.4mg in the media containing culture supernatant of Streptococcus salivarius 119 in BHIS broth. 4. The viable cells of Streptococcus oralis and Streptococcus salivarius 119 were $4.8\times10^7\;and\;7.5\times10^8$ per ml respectively after each culture, wheras being $4.2\times10^7\;and\;5.8\times10^7$ per ml in the combined culture of Streptococcus oralis and Streptococcus salivarius 119. 5. The polymer produced by Streptococcus salivarius 119 was glucan on the thin layer chromatography. 6. The glucan produced by Streptococcus salivarius 119 was water-soluble glucan containing $1\rightarrow6$ linkages as the main linkage on the thin layer chromatography. These results suggested that isolated Streptococcus salivarius 119 inhibited the formation of artificial plaque by the production of water-soluble glucan.

• Asian-Australasian Journal of Animal Sciences
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• v.15 no.12
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• pp.1798-1807
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• 2002

This study was conducted to investigate lactobacillus salivarius subsp. salivarius having probiotic properties to be used as the health adjuncts with fermented milk products. Acid- and bile-tolerant lactobacillus salivarius subsp. salivarius was isolated with lactobacilli MRS broth from faeces of 80 healthy persons (infants, children and adults). It was used as a probiotic strain in fermented milk products. The pH of fermented milk decreased from pH 6.7 to 5.0 and titratable acidity increased from 0.3% to 1.0% by L. salivarius subsp. salivarius (isolation strain 20, 35, and 37), when incubated for 36 h at 37$^{\circ}C$. The number of viable cell counts of fermented milk was maximized at this incubation condition. The SDS-PAGE evidenced no significant change of casein but distinct changes of whey protein were observed by isolated L. salivarius subsp. salivarius for titratable acidity being incubated by 0.9-1.0% at 37$^{\circ}C$. All of the strains produced 83.43 to 131.96 mM of lactic acid and 5.39 to 26.85 mM of isobutyric acid in fermented products. The in vitro culture experiment was performed to evaluate ability to reduce cholesterol levels and antimicrobial activity in the growth medium. The selected L. salivarius subsp. salivarius reduced 23-38% of cholesterol content in lactobacilli MRS broth during bacterial growth for 24 h at 37$^{\circ}C$. All of the isolated L. salivarius subsp. salivarius had an excellent antibacterial activity with 15-25 mm of inhibition zone to E. coli KCTC1039, S. enteritidis KCCM3313, S. typhimurium M-15, and S. typhimurium KCCM40253 when its pH had not been adjusted. Also, all of the isolated L. salivarius subsp. salivarius had partial inhibition zone to E. coli KCTC1039, E. coli KCTC0115 and S. enteritidis KCCM3313 when it had been adjusted to pH 5.7. The selected strains were determined to have resistances of twelve antibiotic. Strains 27 and 35 among the L. salivarius subsp. salivarius showed the highest resistance to the antibiotics. These results indicated that some of the L. salivarius subsp. salivarius (strain 27 and 35) are considered as effective probiotic strains with a potential for industrial applications, but the further study is needed to establish their use as probiotics in vivo.

• Food Science of Animal Resources
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• v.25 no.4
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• pp.483-493
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• 2005

This experiment was carried out to examine the fermentation properties of yogurt prepared with bovine milk and soybean milk at the mixed ratios of 3:1, 2:1, 1:1, 1:2 and 1:3. The effect of bovine milk and soybean milk on promoting the fermentation was higher un pH was $3.75\~4.16$ when Lactobacillus salivarius ssp. salivarius CNU27, lactic culture 1(Lactobacillus delbrueckii ssp. bulgaricus(LB12)), Streptococcus salivarius ssp. thermophilus (ST36) and Lactobacillus acidophilus KCTC3150 were used. Titratable acidity was the highest when lactic culture 1[Lactobacillus delbrueckii ssp. bulgaricus(LB12), Streptococcus salivarius ssp. thermophilus(ST36)] was mea and the mixed ratio of bovine milk and soybean milk was 2:1. The average viable counts of lactic acid bacteria was the highest level of $2.17\times10^9$ cfu/ml when Lactobacillus salivarius ssp. salivarius CNU27 was used, and the mixed ratio of bovine milk and soybean milk was 1:3. the highest lactic acid production was 412.52mM when lactic culture 1[Lactobacillus delbrueckii ssp. bulgaricus (LB12), Streptococcus salivarius ssp. thermophilus (ST36)] was used, and the mixed ratio of bovine milk and soybean milk was 1:1. The production of acetic acid was the highest and the concentration was 394.01mM when lactic culture 2(Bifidobacterium longum, Lactobacillus acidophilus, Streptococcus salivarius ssp. thermophilus) was used and the mixed ratio of bovine milk and soy bean milk was 3:1. Tn the carbohydrate hydrolysis, stachyose was hydrolyzed $53.92\%$ as compared with the control when Lactobacillus salivarius subsp salivarius CNU27 was used, and the mixed ratio of bovine milk and soy bean milk was 1:3. The highest viscosity of yogurt was $1,300\~1,660$ cP when the mixed ratio of bovine milk and soybean milk was 1:3. The overall acceptability, $4.17\pm0.69$, was the highest when Lactobacillus acidophilus KCTC3150 was used and when the mixed ratio of bovine milk and soybean milk was 2:1.

• Proceedings of the Korean Society for Food Science of Animal Resources Conference
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• 2004.05a
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• pp.89-119
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• 2004

This study was conducted to isolate lactobacilli having probiotic characteristics to be used as health adjuncts with fermented milk products. Acid tolerant strains were selected in Lactobacilli MRS broth adjusted to pH 4.0 from 80 healthy persons (infants, children and adults). And bile tolerant strains were examined in Lactobacilli MRS broth in which 1.0% bile salt was added. By estimation above characteristics, the strains No. 27, which was isolated from adult feces, was selected and identified as Lactobacillus salivarius subsp. salivarius based on carbohydrate fermentation and 16S rDNA sequencing. It was used as a probiotic strain in fermented milk products. The pH of fermented milk decreased from pH 6.7 to 5.0 and titratable acidity increased from 0.3% to 1.0% by L. salivarius subsp. salivarius (isolation strain 20, 35, and 37), when incubated for 36 h at $37^{\circ}C$. The number of viable cell counts of fermented milk was maximized at this incubation condition. The SDS-PAGE evidenced no significant change of casein but distinct changes of whey protein were observed by isolated L. salivarius subsp. salivarius for titratable acidity being incubated by $0.9{\sim}1.0%$ at $37^{\circ}C$. All of the strains produced 83.43 to 131.96 mM of lactic acid and 5.39 to 26.85 mM of isobutyric acid in fermented products. The in vitro culture experiment was performed to evaluate ability to reduce cholesterol levels and antimicrobial activity in the growth medium. The selected L. salivarius subsp. salivarius reduced $23{\sim}38%$ of cholesterol content in lactobacilli MRS broth during bacterial growth for 24 hours at $37^{\circ}C$. All of the isolated L. salivarius subsp. salivarius had an excellent antibacterial activity with $15{\sim}25$ mm of inhibition zone to E. coli KCTC1039, S. enteritidis KCCM3313, S. typhimurium M-15, and S. typhimurium KCCM40253 when its pH had not been adjusted. Also, all of the isolated L. salivarius subsp. salivarius had partial inhibition zone to E. coli KCTC1039, E. coli KCTC0115 and S. enteritidis KCCM3313 when it had been adjusted to pH 5.7. The selected strains were determined to have resistances of twelve antibiotic. Strains 27 and 35 among the L. salivarius subsp. salivarius showed the highest resistance to the antibiotics. Purified ${\alpha}$-galactosidase was obtained by DEAE-Sephadex A-50 ion exchange chromatography, Mono-Q ion exchange chromatography and HPLC column chromatography from L. salivarius subsp. salivarius 27. The specific activity of the purified enzyme was 8,994 units/mg protein, representing an 17.09 folds purification of the original cell crude extract. The molecular weight of enzyme was identified about 53,000 dalton by 12% SDS-PAGE. Optimal temperature and pH for activity of this enzyme were $40^{\circ}C$ and 7.0 respectively. The enzyme was found to be stable between 25 and $50^{\circ}C$. ${\alpha}$-galactosidase activity was lost rapidly below pH 5.0 and above pH 9.0. This enzyme was liberated galactose from melibiose, raffinose, and stachyose, and also the hydrolysis rate of substrate was compound by HPLC. These results indicated that some of the L. salivarius subsp. salivarius (strain 27 and 35) are considered as effective probiotic strains with a potential for industrial applications, but the further study is needed to establish their use as probiotics in vivo.

• Journal of Oral Medicine and Pain
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• v.34 no.2
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• pp.153-167
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• 2009

The present study was performed to observe the effect of phytoncide on oral normal microflora and the inhibitory effect of the surviving resident oral bacteria on Pr. intermedia. In this study, saliva from each of 20 healthy subjects was treated with 1% phytoncide from Japanese Hinoki (Chamaecyparis obtusa Sieb. et Zucc.). Surviving salivary bacteria were isolated on blood agar plates and identified by 16S rDNA sequencing. In order to select inhibitory isolates against Pr. intermedia, the isolates from the phytoncide-treated saliva were cultured with Pr. intermedia. The results were as follows: 1. Among the 200 surviving resident oral bacterium, 148(74.0%) bacterium inhibit the growth of Pr. intermedia on blood agar plates. 2. The 200 surviving resident oral bacterium were 109 Streptococcus salivarius(54.5%), 25 Streptococcus sanguinis(12.5%), 15 Streptococcus mitis(7.5%). 3. Among the 148 bacteria which inhibit Pr. intermedia, Streptococcus salivarius was 85.3%(93/109), Streptococcus sanguinis was 64.0%.(16/25), Streptococcus mitis was 54.3%(8/15), Streptococcus parasanguinis was 66.7%(6/9), and Streptococcus Alactolyticus was 100%(8/8). Taken together, among the surviving resident oral bacterium, Streptococcus salivarius, Streptococcus sanguinis, Streptococcus mitis were mainly observed to inhibit Pr. intermedia. and they may exert an additional inhibitory activity against the periodontopathic bacterium. Therefore, phytoncide can be used for preventing and ceasing the progress of periodontal disease and halitosis, and thus is expect to promote oral health.

• Journal of Microbiology
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• v.43 no.2
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• pp.204-208
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• 2005

The objective of this study was to both isolate and identify non-mutans streptococci organisms (non­MSO) from dental plaques recovered on mitis-salivarius sucrose bacitracin agar (MSB) plates. The dental plaque samples, which had been collected from 63 human subjects, were diluted and plated on MSB. The bacteria growing on the MSB plates were then identified with biochemical tests, as well as with 16S rDNA cloning and sequencing techniques. Our data indicated that bacteria from 30 subjects had been recovered on the MSB plates. Among the 21 typical colonies selected from the 30 subjects, 12 colonies, derived from 10 subjects, were identified as non-MSO. These 12 colonies were determined to be Streptococcus anginosus (8 colonies), S. sanguinis (1 colony), and Pantoea agglomerans (3 colonies). These results strongly suggest that a new selective medium will be required for the reliable isolation of mutans streptococci.

• Journal of Korean society of Dental Hygiene
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• v.12 no.1
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• pp.81-91
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• 2012

Objectives : By investigating antimicrobial activity of natural extracts, identifying its usefulness as antibiotic material to oral bacteria. Methods : Antimicrobial activity tests of 25 natural extracts were implemented on 6 type strains alleged to cause dental caries and 10 clinical strains isolated and identified from dental caries. Results : Among medicinal plants, Coptis japonica, Scutellaria baicalensis, Dictamnus dasycarpus showed antimicrobial activity. Among lichens, 3 methanol extracts and 6 acetone extracts showed antimicrobial activity. Minimum Inhibitory Concentration(MIC) test of Usnea aurantiacoatra KoLRI 004184, an acetone extract of lichen with the highest antimicrobial activity, on 6 type strains involved in dental caries and 10 strains isolated from dental caries resulted as follows; Actinomyces oris 1041 $12{\mu}g/ml$, for Corynebacterium durum 3151 $13{\mu}g/ml$, for Rothia dentocariosa KCTC $3204^T$ $14{\mu}g/ml$, for R. dentocariosa 911 $15{\mu}g/ml$, for R. dentocariosa 1822 $12{\mu}g/ml$, for Lacto-bacillus casei KACC $12413^T$ $12{\mu}g/ml$, for L. acidophilus KACC $12419^T$ $16{\mu}g/ml$, for L. rhamnosus 2421 $15{\mu}g/ml$, for Streptococcus mutans KCTC $3065^T$ $15{\mu}g/ml$, for S. mutans 121 $13{\mu}g/ml$, for Streptococcus oralis 2221 $18{\mu}g/ml$, for S. salivarius KCTC 5512 $14{\mu}g/ml$, for S. salivarius 122 $12{\mu}g/ml$, for S. sanguinis KCTC $3284^T$ $14{\mu}g/ml$, for S. sanguinis 912 $14{\mu}g/ml$, for Neisseria sp. KEM232 $12{\mu}g/ml$. Conclusions : Even a small amount of extract from lichen Usnea aurantiacoatra KoLRI 004184 showed very high level of antimicrobial activation against all 16 bacterial strains involved in dental caries.