Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
  • A61K35/66—Microorganisms or materials therefrom
  • A61K35/74—Bacteria
  • A61K35/741—Probiotics
  • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23G—COCOA; COCOA PRODUCTS, e.g. CHOCOLATE; SUBSTITUTES FOR COCOA OR COCOA PRODUCTS; CONFECTIONERY; CHEWING GUM; ICE-CREAM; PREPARATION THEREOF
  • A23G4/00—Chewing gum
  • A23G4/06—Chewing gum characterised by the composition containing organic or inorganic compounds
  • A23G4/12—Chewing gum characterised by the composition containing organic or inorganic compounds containing microorganisms or enzymes; containing paramedical or dietetical agents, e.g. vitamins
  • A23G4/123—Chewing gum characterised by the composition containing organic or inorganic compounds containing microorganisms or enzymes; containing paramedical or dietetical agents, e.g. vitamins containing microorganisms, enzymes
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; PREPARATION THEREOF
  • A23C9/00—Milk preparations; Milk powder or milk powder preparations
  • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
  • A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; PREPARATION THEREOF
  • A23C9/00—Milk preparations; Milk powder or milk powder preparations
  • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
  • A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
  • A23C9/1236—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt using Leuconostoc, Pediococcus or Streptococcus sp. other than Streptococcus Thermophilus; Artificial sour buttermilk in general
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; PREPARATION THEREOF
  • A23C9/00—Milk preparations; Milk powder or milk powder preparations
  • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
  • A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
  • A23C9/1238—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt using specific L. bulgaricus or S. thermophilus microorganisms; using entrapped or encapsulated yoghurt bacteria; Physical or chemical treatment of L. bulgaricus or S. thermophilus cultures; Fermentation only with L. bulgaricus or only with S. thermophilus
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23K—FODDER
  • A23K10/00—Animal feeding-stuffs
  • A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
  • A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
  • A23K10/18—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions of live microorganisms
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23K—FODDER
  • A23K40/00—Shaping or working-up of animal feeding-stuffs
  • A23K40/30—Shaping or working-up of animal feeding-stuffs by encapsulating; by coating
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23K—FODDER
  • A23K50/00—Feeding-stuffs specially adapted for particular animals
  • A23K50/40—Feeding-stuffs specially adapted for particular animals for carnivorous animals, e.g. cats or dogs
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K8/00—Cosmetics or similar toiletry preparations
  • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
  • A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
  • A61K8/99—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from microorganisms other than algae or fungi, e.g. protozoa or bacteria
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
  • A61P1/02—Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/04—Antibacterial agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
  • A61Q11/00—Preparations for care of the teeth, of the oral cavity or of dentures; Dentifrices, e.g. toothpastes; Mouth rinses
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; PREPARATION THEREOF
  • A23C2220/00—Biochemical treatment
  • A23C2220/20—Treatment with microorganisms
  • A23C2220/206—Slime forming bacteria; Exopolysaccharide or thickener producing bacteria, ropy cultures, so-called filant strains
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
  • A23V2400/00—Lactic or propionic acid bacteria
  • A23V2400/21—Streptococcus, lactococcus
  • A23V2400/249—Thermophilus
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
  • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10S435/00—Chemistry: molecular biology and microbiology
  • Y10S435/8215—Microorganisms
  • Y10S435/822—Microorganisms using bacteria or actinomycetales
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
  • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10S435/00—Chemistry: molecular biology and microbiology
  • Y10S435/8215—Microorganisms
  • Y10S435/822—Microorganisms using bacteria or actinomycetales
  • Y10S435/885—Streptococcus

Definitions

• the present invention relates to the incorporation in the oral microflora of exogenous lactic bacteria which are able to modulate the colonization of A. naeslundii and to reduce the severity of A. naeslundii-r&ldited diseases.
• the mouth (oral cavity) contains a resident and a non-resident microflora.
• the first includes microorganisms that are able to establish a more or less permanent residence on the oral surfaces. These bacteria are mainly localised on the tongue, the buccal mucosa and the teeth while the gingiva, lips, cheeks, palate and floor of the mouth only support a very sparse microflora.
• the dental plaque is a film that forms on the surface of teeth consisting of bacterial cells in a matrix of extracellular polysaccharides and salivary products. Immediately after eruption, the teeth are covered with an amorphous layer of saliva, the acquired enamel pellicle (AEP) that is about 1.3 ⁇ m thick and cannot be removed by normal tooth brushing.
• AEP acquired enamel pellicle
• the deposition of bacteria on teeth follows immediately the formation of the AEP and plaque becomes evident in 8- 12 hours as a multi-layered structure.
• the first layer consists of bacteria (earliest colonisers) that attach to teeth mainly via specific adhesin-receptor recognition; it forms a substratum for the second colonisers that adhere one to the other via analogous specific binding or via simple juxtaposition.
• the natural resident microflora includes microorganisms selected from Streptococcus, Veillonella, Bacteroides and Haemophilus.
• Streptococci and Actinomyces predominate but a variety of Gram positive and negative cocci and rods can be found.
• Actinomyces naeslundii genospecies 1 (formerly A. naeslundi ⁇ ) and 2 (formerly A. viscosus) are common members of human dental plaque. They are among the strongest plaque forming oral strains, because of their capacity to firmly adhere to the teeth and to coaggregate with many other bacterial species, thus fostering their establishment in the mouth.
• the transient microflora comprises exogenous bacteria that can be occasionally present in the mouth, but that do not establish a permanent residence
• All the food bacteria, and in particular lactic acid bacteria, can be part of this transient microflora.
• lactic bacteria strains that are not part of the resident microflora of the mouth, that are low acidifying and that are capable of adhering directly to the pellicle of the teeth.
• These bacteria are particularly used for treating or preventing dental caries and periodontal infection that are caused by cariogenic microorganism such as Streptococcus mutans and Streptococcus sobrinus.
• Exogenous bacteria can also produce factors that inhibit the growth of the resident microflora in the mouth.
• EP759469 Societe des Produits Nestle
• bacteriocins produced by Micrococcus varians for inhibiting the development of the oral pathogens S. sobrinus, S. sanguis, S. mutans and A. viscosus.
• the application of bacteriocins is also one of the investigated strategies which have been set up to reduce tooth caries. These molecules have attracted interest as prospective anticaries agents and as factors important in modulating colonisation of the oral cavity.
• the present invention aims to provide the use of lactic bacteria that are exogenous to the oral microflora, which have been selected for their ability to adhere to the tooh surface and to produce a growth inhibition factor, for the preparation of a composition intended for reducing dental plaque and for treating or preventing root caries or other disease related to Actinomyces naeslundii in mammals.
• the lactic bacteria may be selected from the group consisting of Streptococcus thermophilus, Lactococcus lactis subsp. lactis, and Lactococcus lactis subsp. lactis biovar diacetylactis and particularly from the group consisting of the strains CNCM 1-1984, CNCM 1-1985, CNCM 1-1986, CNCM 1-1987.
• Another object is to provide a composition for maintaining the health of the mouth by reducing the colonization of Actinomyces naeslundi, said composition comprises an exogenous lactic bacteria that has been selected for its ability to adhere to the tooh surface and to produce a growth inhibition factor.
• Such a composition may contain at least 10 4 -10 9 cfu /g of lactic bacteria.
• the invention also provides a method for the prevention or the treatment of Actinomyces naeslundi related infections, particularly dental plaque extent and root caries in a mammal, comprising the step of feeding a mammal a composition containing at least one lactic bacteria strain selected for its ability to adhere to the tooh surface and to produce a growth inhibition factor, said composition reduces the colonization of Actinomyces naeslundi.
• the mouth defines the oral cavity of humans or animals such as pets, composed by the oral mucosa (gums, lips, cheeks, palate and floor of the mouth), the tongue and the teeth (including artificial structures).
• inhibitor growth factor defines any extracellular substance produced by the adherent exogenous lactic bacteria that enables it to inhibit the growth of A.naeslundi.
• the use of an exogenous lactic bacteria that has been selected for its ability to adhere to the tooh surface and to produce a growth inhibition factor, for the preparation of a composition intended for reducing dental plaque and for treating or preventing root caries or other disease related to Actinomyces naeslundi, is concerned.
• the lactic bacteria may be selected from the group consisting of
• Streptococcus thermophilus Lactococcus lactis subsp. lactis, and Lactococcus lactis subsp. lactis biovar diacetylactis and particularly from the group consisting of the strains Sreptococcus thermophilus (NCC 1529) (CNCM 1-1984),
• the lactic bacteria is preferably from dairy origin (i.e. originating from milk or cheese, for example).
• the lactic bacteria according to the invention is "low acidifying", which means that it is less acidifying than pathogenic strains. Accordingly, it can contribute to a pH in the oral cavity of about 5.5-7.
• strains have been selected among latic bacteria strains for their capacity of adherence to the pellicle of the teeth, their optimal growth temperature is about 37°C, which is the temperature in the oral cavity. They are also capable of producing a growth inhibition factor, which combined to their adhesion properties allow them to significantly decrease the colonization extent of A. naeslundii genospecies 1 and 2.
• At least one lactic bacteria strain in combination with a bacteriocin, for example.
• the lactic bacteria strains may be included in a food, pet food, cosmetic or pharmaceutical composition, for example. Accordingly, these compositions are preferably toothpaste, mouth rinse, gum, spray, beverage, candies, infant formula, ice cream, frozen dessert, sweet salad dressing, milk preparations, cheese, quark, yogurt, acidified milk, coffee cream or whipped cream, for example.
• the exogenous lactic bacteria may be used in an amount of at least 10 4 - 10 9 cfu /g of lactic bacteria.
• the effect of incorporating the above-mentioned bacteria in the oral microflora was tested in a rat model.
• the strains CNCM 1-1985 and CNCM- 1986 were able to modulate the oral microbial ecology, significatively reducing the number of total CFU. More specifically, the strains were able to significantly decrease the colonization extent of A. naeslundii genospecies 2, with which the rats had been infected (see examples).
• Fermentation patterns 49 simple sugars were tested with the api 50 CH bioMerieux strip test (bioMerieux SA, 69280 Marcy-l'Etoile, France) and the results are given in Table 1.
• strains Sreptococcus thermophilus (NCC 1529), Sreptococcus thermophilus (NCC 1561), Lactococcus. lactis subsp. lactis (NCC 2211), Lactococcus. lactis subsp. lactis biovar dioacetylactis (NCC 2225) were deposited under the Budapest Treaty, at the Collection Nationale de Culture de Microorganismes (CNCM 1-1984, CNCM 1-1985, CNCM 1-1986 and CNCM I- 1987 respectively), 25 rue du dondel Roux, 75724 Paris, France, on March 3 rd , 1998.
• the second main object of the present invention relates to a composition for maintaining the health of the mouth by reducing the colonization of A. naeslundii in mammals, said composition comprises an exogenous lactic bacteria, which has been selected for its ability to adhere to the tooh surface and to produce a growth inhibition factor.
• compositions are particularly intended for the prophylaxis or the treatment of dental plaque and infection related to A. naeslundii disease such as root caries, for example.
• the lactic bacteria strain according to the present invention is selected from the group consisting of Streptococcus thermophilus, Lactococcus lactis subsp. lactis, and Lactococcus lactis subsp. lactis biovar diacetylactis and preferably from the group consisting of the strains CNCM 1-1984, CNCM I-
• Such a ; composition may contain at least 10 4 -10 9 cfu /g of lactic bacteria.
• Synergistic compositions may also be prepared, adding at least one bacteriocin, which is active against Gram-positive oral bacteria.
• the oral hygiene compositions may comprise 0.00001 to 50%, and preferably from 0.00001 to 15% of purified bacteriocin, by weight of the composition.
• the bacteriocin is preferably variacin (EP 0 759 469).
• an oil-soluble antioxidant may also be included. Suitable antioxidants include the “tocopherols", butyl-hydroxyanisole (BHA), butyl-hydrxytoluene (BHT), and ascorbyl palmitate.
• BHA butyl-hydroxyanisole
• BHT butyl-hydrxytoluene
• ascorbyl palmitate ascorbyl palmitate.
• the oil soluble antioxidant is present in amounts of from
• Suitable abrasives for use in dentifrice compositions of the present invention include calcium carbonate, calcium aluminosilicate, alumina, hydrates alumina, zinc orthophosphate, plastic particles, and silica, of which silica is the preferred abrasive.
• compositions according to the invention will have a pH which is orally acceptable and within which the activity of the said lactic bacteria is not compromised.
• the pH may be in the range 3.0-9.5, preferably in the range 3.5 to 6.5.
• These compositions may be prepared by conventional processes comprising admixing the ingredients together in the appropriate relative amounts and finally, and if necessary, adjusting the pH to desired value.
• Actinomyces naeslundii genospecies 1 (formerly A. naeslundii) and 2 (formerly A. viscosus) are among the strongest plaque forming oral strains. They are commonly isolated at root caries sites, in particular in humans over 40 years, and they are believed to be the major aetiological agent of this disease.
• the invention also provides a method for the prevention or the treatment of Actinomyces naeslundi-r&lated infections in mammals, particularly dental plaque extent and root caries, comprising the step of feeding the mammal a composition containing at least one lactic bacteria strain selected for its ability to adhere to the tooh surface and to produce a growth inhibition factor.
• the amount to be administred may be of at least about 10 4 -10 9 cfu /g of lactic bacteria.
• L. lactis NCC2211 The strains S. thermophilus NCC 1561 (CNCM 1-1985) and L. lactis subsp. lactis NCC2211 (CNCM 1-1986) (hereinafter L. lactis NCC2211) were incorporated in vitro in a biofilm mimicking dental plaque in vitro.
• the two selected strains L. lactis NCC2211 and S. thermophilus NCC1561 were inoculated in an in vitro system in which a biofilm, composed by bacteria commonly found in the human mouth after 40 years, developed on saliva coated-hydroxyapatite discs.
• Fluid Universal Medium FUM
• the assays were done in triplicate and the mixtures with and without the dairy strains were tested in parallel.
• the strains listed in the Table 2 were used.
• Saliva pellicle formation cover synthetic hydroxyapatite discs of 10 mm diameter (HY-APATITE ® , Euro-Crystals, Landgraaf, The Netherlands) with 800 ⁇ l of human saliva and incubate for 4 h at room temperature under shaking (1 disc/well in a 24 holes sterile Nunclon plate).
• Bacterial consortium preparation grow S. thermophilus NCC1561, L. lactis subsp. lactis NCC2211, S. sobrinus OMZ176, S. oralis OMZ607, A. naeslundii OMZ745, V. dispar OMZ493 and F.
• nucleatum OMZ596 overnight at 37°C in anaerobiosis in FUM-glucose (S. thermophilus NCC1561 in FUM-lactose), adjust the OD 550 to 1 with FUM and pool 2 ml of each oral bacterial suspension with 2 ml of either S. thermophilus NCC1561 or L. lactis subsp. lactis NCC2211.
• the control mixture contains the five oral strains only.
• thermophilus NCC 1561 and L. lactis NCC2211 could be incorporated and grown in the plaque-like biofilm on the S-HA discs, and their total CFU/disc after 40.5 h are given in the Table 4.
• N 3. *: p-values are calculated in respect to the control (*: p ⁇ 0.05; **: p ⁇ 0.01); °: p- values are calculated in respect to the "+NCC1561" treatment (°: ⁇ 0.05; °°: p ⁇ 0.01).
• N 3. *: p-values are calculated in respect to the control (*: p ⁇ 0.05; **: p ⁇ 0.01); °: p- values are calculated in respect to the "+NCC2211" treatment (°: p ⁇ 0.05; °°: p ⁇ 0.01).
• S. thermophilus NCC1561 and L. lactis NCC2211 could be incorporated in a biofilm mimicking dental plaque and were able to modulate the oral microflora, significantly reducing the number total of cfu, and more specifically, these strains were able to significantly decrease the colonization extent of A. naeslundii genospecies 2. In addition the strains could inhibit the growth of A. naeslundii genospecies 1 and 2 in co-cultures.
• Example 2 in- vivo trials.
• the cariogenic strains were associated on days 21 and 22, while association of the dairy strains started on day 23 and lasted until day 57.
• the animals were fed the dairy strains as supplement in a yogurt base that was included in the normal diet, as explained in the section.
• the rats teeth were swabbed at the end of the study, on day 58.
• the rats On day 20 the dams were removed, and the rats started to be fed ad libitum with the modified diet 2000a containing 40% sucrose, 28% substitute for skim milk (soya protein extract SVPRO-PP 1611 39.4%, lactose 49.3%, 0.6%, L- Methionine 0.3%, L-Lysine HC1 0.1%), 24% wheat flour, 5% brewer's yeast, 2% Gevral ® Instant Protein (Whitehall-Robins SA, 6301 Switzerland) and 1% NaCl. During the association period (days 21 and 22) the drinking water was supplemented with 2% glucose and 2% sucrose to support the implantation of the associated bacteria.
• test diet consisted of 18 yogurt meals containing the test strains alternating with 18 meals of the modified diet 2000a previously described.
• Bacterial strains The strains listed in the Table 8 were used.
• mice were arranged in 3 treatments (Table 9). Each treatment consisted of 10 pups that were distributed in 1 per cage.
• the tested LAB strains were associated daily (since contained in the yogurt base meal), starting on day 23. 2 frozen vials, each containing 2xlO ⁇ viable cells of the test strain, were mixed in 200 ml of yogurt in order to have at least 10 9 CFU/ml. S. thermophilus NCC1536, a non S-HA adhering strain, was used as a negative control.
• the yogurt and diet 2000a meals were offered alternatively 18 times per day at 20-min intervals (Table 10). Therefore, each animal received in total 18 ml of yogurt and 7.2 g of powdered diet.
• the meals were dispensed in the food cups of a programmed feeder machine that automatically offered to the animals the right meal at the exact time.
• mice Five rats per treatment were swabbed on day 58.
• the swab suspensions were either plated on Petri dishes for CFU (colony forming units) counts or immobilized on glass slides for immunofluorescence for TCN (total cells number) counts.
• the percentages of A. viscosus OMZ105 on the total CFU did not decrease in parallel, therefore one can deduce that the growth antagonism effect was also displayed versus other species, i.e. Veillonellae, and consequently it is a global effect which is observed.
• strains CNCM 1-1985 and CNCM- 1986 are able to modulate the oral microbial ecology, significatively decreasing the colonization extent of A. naeslundii genospecies 2, with which the rats had been infected.
• Example 3 Production and initial analysis of surfactant substances from S. thermophilus NCC1561 and S. thermophilus NCC1536.
• thermophilus NCC1561 and S. thermophilus NCC1536 were grown overnight in 1 1 of Belliker at 42 °C.
• the procedure described in Busscher et al. (1997) Appl. Environ. Microbiol. 63, 3810-3817, (Busscher et al., 1997) was used.
• the crude biosurfactant suspension was first analyzed by SDS-PAGE and then submitted to surface tension measurements.
• S. thermophilus NCC1561 and S. thermophilus NCC1536 cells are able to release substances with a surfactant activity. It is therefore possible that the biosurfactant produced by S. thermophilus NCC 1561 makes the bacterium itself and the other oral strains established close to it detach from the tooth surface. By contrast, this action would not be displayed by S. thermophilus NCC 1536, since this strain does not adhere to the teeth.
• Toothpaste is prepared by adding 10 5 cfu/ml of at least one of the lactic bacteria strain CNCM 1-1984, CNCM 1-1985, CNCM 1-1986, CNCM 1-1987 in a lyophilised form, to the following mixture containing: 1.65% Cetyl pyridinium chloride, 33.0% Sorbitol (70% soln), 25.0% Glycerin, 2.0% Sodium carboxymethyl cellulose, 0.25% Sodium fluoride, 26.3% Silica (RP 93), 8.1% Thickening Silica (Sident 22), 0.5% Sodium saccharine, 3.2% Poloxamer (Pluronic F108).
• This toothpaste is intended for the prophylaxis or the treatment of root caries, dental plaque and other infections induced by A.naeslundii species.
• 5 1 MRS culture medium are sterilised for 15 min at 121°C and then inoculated with 5% by volume of an active culture of at least one of the S. thermophilus strain CNCM 1-1984, CNCM 1-1985 containing approximately 10 cfu/ml. After incubation for 8 h at 41°C, a starter containing 4.5.10 cfu/ml is obtained.
• the yoghurt obtained has a good firm and smooth texture and is intended for the health of the mouth.
• a chewing gum for preventing or treating root caries, dental plaque or other A.naeslundii- ⁇ ela.ted deseases can be prepared adding an active culture of at least one of the S. thermophilus strain CNCM 1-1984, CNCM 1-1985 so that it contains approximately 10 4 to 10 9 cfu/g, to the following typical ingredients: 67.5 % Xylitol, 20 % Gum base, 5 % Calcium carbonate, 3 % Glycerin, 2 % Pluronic F127, 1 % Cellulose gum, 0.5 % Balast compounds and 1 % Flavor.
• a pet food for mouth health is obtained by preparing a feed mixture made up of corn, corn gluten chicken and fish meal, salts, vitamins and minerals.
• the feed mixture is fed into a preconditioner and moistened.
• the moistened feed leaving the preconditioner is then fed into an extruder-cooker and gelatinised.
• the gelatinised matrix leaving the extruder is forced through a die and extruded.
• the extrudate is cut into pieces suitable for feeding to dogs, dried at about 110°C for about 20 minutes and cooled to form pellets which have a water activity of about 0.6.
• the pellets are sprayed with 3 coating mixtures.
• Each coating mixture contains active culture of at least one of the S.
• the pellets contain approximately 10 4 to 10 9 cfu/g of said strains.

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