WO2005009139A1 - Produit d'animaux d'elevage contenant une bacterie probiotique enterococcus - Google Patents

Produit d'animaux d'elevage contenant une bacterie probiotique enterococcus Download PDF

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Publication number
WO2005009139A1
WO2005009139A1 PCT/DK2004/000519 DK2004000519W WO2005009139A1 WO 2005009139 A1 WO2005009139 A1 WO 2005009139A1 DK 2004000519 W DK2004000519 W DK 2004000519W WO 2005009139 A1 WO2005009139 A1 WO 2005009139A1
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WIPO (PCT)
Prior art keywords
product
farm animal
farm
bacteria
cfu
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PCT/DK2004/000519
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English (en)
Inventor
Jane A. Z. Leedle
Steven C. Johnson
William P. Kautz
Kelly F. Lechtenberg
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Chr. Hansen A/S
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Priority claimed from EP03077658A external-priority patent/EP1510135A1/fr
Application filed by Chr. Hansen A/S filed Critical Chr. Hansen A/S
Publication of WO2005009139A1 publication Critical patent/WO2005009139A1/fr

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    • 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
    • A23K50/30—Feeding-stuffs specially adapted for particular animals for swines
    • 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
    • A23K50/00—Feeding-stuffs specially adapted for particular animals
    • A23K50/10—Feeding-stuffs specially adapted for particular animals for ruminants
    • 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
    • A23K50/70—Feeding-stuffs specially adapted for particular animals for birds
    • A23K50/75—Feeding-stuffs specially adapted for particular animals for birds for poultry

Definitions

  • the present invention relates to a farm animal product comprising probiotic Enterococcus bacteria and the use of this product to reduce the number of pathogenic Escherichia coli O157:H7 cells in farm animals such as cattle.
  • the animal feed industry such as the beef cattle industry
  • challenges like never before and one of the most critical challenges to the industry is food safety.
  • the consumer and governmental agencies are requiring that beef sold in restaurants, grocery stores and meat markets be as safe and pathogen free as possible.
  • Meat packing companies are looking to the feed yards and the cattle producers to implement strategies to help achieve this goal.
  • E. coli O157:H7 is one of hundreds of strains of the bacterium. Although most strains are harmless and live in the intestines of healthy humans and animals, this strain produces a powerful toxin and can cause severe illness. It also possesses other significant attributes, which contribute to its ability to cause disease. One of the more notable of its characteristics is the size of the infectious dose, which is incredibly small in comparison with those for most other food-borne pathogens. Figures as low as two bacteria per 25 g food have been quoted capable of creating a disease condition.
  • the strain E. coli O157:H7 was first recognized as a cause of illness in 1982 during an outbreak of severe bloody diarrhea; the outbreak was traced to contaminated hamburgers. Since then, most infections have resulted from eating undercooked ground beef.
  • the combination of letters and numbers in the name of the bacterium refers to the specific markers found on its surface and distinguishes it from other types of E. coli.
  • Another pathogen of concern includes strains of Salmonella, with both E. coli and
  • Salmonella commonly existing in the gastrointestinal tracts of cattle. These organisms are endemic and commonly found in virtually all phases of production. While they may not cause a problem in the host animal they can cause illness and even death in humans. Cattle become “infected” with this organism through exposure in their natural environment. After the organism is ingested it travels to the intestine where it adheres to the tract lining. Meat is "contaminated” by the organism during the slaughtering and processing stages when intestinal contents can come in contact with other meat surfaces and subsequently become mixed with ground beef.
  • an E. coli infection can lead to bloody diarrhea and even kidney failure.
  • the infection can also cause a complication called hemolytic uremic syndrome, in which the red blood cells are destroyed and the kidneys fail.
  • hemolytic uremic syndrome is the principal cause of acute kidney failure in children, and most cases of hemolytic uremic syndrome are caused by the strain E. coli O157:H7.
  • DFM Direct-Fed Microbial
  • DFM Most bacterial-based DFM's are beneficial because they have effects in the lower gut and not in the rumen.
  • Lactobacillus acidophilus produces lactic acid, which may lower the pH in small intestines to levels that inhibit the growth of pathogenic microbes, one of the reasons for the current interest.
  • Early research with DFM in ruminants first involved applications for young calves fed milk, calves being weaned, or cattle being shipped. These animals, in many cases, are highly stressed or had a microbial gut ecosystem that was not fully mature. Young cattle have immature digestive tracts that are obviously more prone to upset by pathogenic bacteria. Cattle that are shipped are often on limited feed and water for prolonged periods of time during transit. During these periods microbial populations may decrease in numbers, thus resulting in digestive tracts that are in less than optimal condition. Large doses of beneficial organisms were thought to re-colonize a stressed intestinal environment and return gut function to normal.
  • Lactobacillus acidophilus BG2FO4 exhibits a high degree of pathogen oriented anti-microbial activity and is an excellent choice for use in beef cattle for this purpose.
  • U.S. Patent No. 5,718,894 (“the '894 patent”) describes a formulation for use in the promotion of growth or weight gain in a farm animal.
  • the '894 patent states that the formulation comprises two groups of bacteria: a so-called first bacterium capable of producing lactic acid in the gastrointestinal tract of the animal; and a second bacterium capable of producing a bactericide to which the bacteria are resistant, wherein said second bacterium is a Bacillus.
  • the '894 patent states that bactericide produced by the Bacillus strain is capable of combating microorganisms that are the positive agent of enteric disorders, e.g. Staphylococcus aureus, E. coli and Salmonella (see column 2, lines 23-31).
  • the '894 patent states that examples of bacterium capable of producing lactic acid are bacteria of the genus Lactobacillus or Enterococcus. According to the '894 patent, they are distinguished by their ability to produce lactic acid and thus reduce the local pH in the gastrointestinal tract of the animal (see column 2, lines 8-22).
  • a specific formulation for use in pigs is described in the '894 patent. It is composed of the four strains Lactobacillus, Enterococcus faecalis, Enterococcus faecium and Bacillus licheniformis. The '894 patent states that each strain is used at 10 9 cfu/g.
  • the problem to be solved by the present invention includes the provision of a composition (also referred to herein as "farm animal product") that has an improved ability with respect to decreasing the number of Escherichia coli O157:H7 in farm animals (preferably cattle) when the farm animals are challenged with Escherichia coli Ol 57:H7 pathogen.
  • a composition also referred to herein as "farm animal product” that has an improved ability with respect to decreasing the number of Escherichia coli O157:H7 in farm animals (preferably cattle) when the farm animals are challenged with Escherichia coli Ol 57:H7 pathogen.
  • compositions comprising Enterococcus strains works better than a corresponding composition comprising similar amounts (CFU/g) ol Lactobacillus acidophilus strains.
  • the farm animal product may comprise smaller amounts of Lactobacillus acidophilus bacteria than those ol Enterococcus bacteria.
  • one aspect of the invention relates to a farm animal product comprising at least 10 CFU/g of probiotic Enterococcus bacteria, characterized in that, if the product contains Lactobacillus acidophilus, the product has at least 2.5 times more ol Enterococcus bacteria than Lactobacillus acidophilus bacteria measured as CFU/g.
  • Another aspect of the invention relates to a method of feeding a farm animal comprising feeding the farm animal with a farm animal product comprising at least 10 6 CFU/g of probiotic Enterococcus bacteria, characterized in that, if the product contains Lactobacillus acidophilus, the product has at least 2.5 times more ol Enterococcus bacteria than Lactobacillus acidophilus bacteria measured as CFU/g.
  • a further aspect of the invention relates to a method of feeding a farm animal comprising feeding the farm animal with a farm animal product comprising at least 10 6 CFU/g of probiotic Enterococcus bacteria, wherein said product reduces the number of Escherichia coli O157:H7 cells quantified in faeces of challenged animals by at least 1.5 logs.
  • the term "probiotic” is a well-defined term in the art and relates to a class of microorganisms defined as live microbial organisms that confer health benefit to farm animal hosts or is not pathogenic to same.
  • the beneficial effects include improvement of the microbial balance of the intestinal micro flora and the improvement of the properties of the indigenous micro flora when, for example, the microorganism is orally administered.
  • Enterococcus is a well-known and well-defined term for this Enterococcus genus of bacteria species. For further details see, e.g., the standard reference book Bergeys Manual of Systematic Bacteriology. Based on the general knowledge of one of ordinary skill in the art, the skilled person is capable of determining whether or not a specific Enterococcus bacterium of interest is a bacterium of the Enterococcus genus.
  • CFU Colony Forming Units.
  • CFU/g refers to CFU/g of farm animal product.
  • the farm animal product comprises suitable farm animal feedstuff ingredients in addition to an Enterococcus bacteria.
  • suitable farm animal feedstuff ingredients may be termed farm animal feedstuff ingredients known per se or farm animal feedstuff ingredients.
  • these ingredients should be in concentrations adjusted to meet animal's dietary requirements and may include nutrient ingredients such as animal protein products, at about 0 - about 95 weight percent; plant protein products, at about 0 - about 95 weight percent; poultry egg products, at about 0 - about 25 weight percent.
  • nutrient ingredients such as animal protein products, at about 0 - about 95 weight percent; plant protein products, at about 0 - about 95 weight percent; poultry egg products, at about 0 - about 25 weight percent.
  • the farm animal product may also comprise other suitable ingredients such as antibiotics such as Sarafin, Romet, Terramycin at about 0.01 - about 50 weight percent; cyanocobalamin at about 40 - about 60 mg/kg; D-biotin at about 5 - about 20 mg/kg; D-pantothenic acid at about 250 - about 350 mg/kg; folic acid at about 10 - about 30 mg/kg; L-ascorbyl-2-polyphosphate (STAY- C, stable form of vitamin C) at about 1,000 - about 4,000 mg/kg; myo-inositol at about 3,000 - about 4,000 mg/kg; niacin at about 600 - about 800 mg/kg; p-amino-benzoic acid at about 350 - about 450 mg/kg; pyridoxine hydrochloride at about 40 - about 60 mg/kg; riboflavin at about 125 - about 175 mg/kg; thiamine hydroch
  • antibiotics such as Sarafin, Romet
  • the farm animal product may be present in any suitable form, such as a powder, liquid or in form of pellets or tablets.
  • One embodiment of a farm animal product is a composition comprising Enterococcus bacteria in a bolus, or, in a further embodiment, in a gelatin bolus.
  • a particular embodiment of a farm animal product is a composition comprising Enterococcus bacteria, Glucidex IT12 (about 30%) and Type 4A Act Molecular Sieve Powder (about 10%).
  • the farm animal product may be in form of, e.g., two different compositions.
  • Such an embodiment includes one composition comprising the suitable farm animal feedstuff ingredients and the other composition comprising the Enterococcus bacteria as described herein.
  • the farm animal product is comprised of such two compositions and accompanied by suitable instructions to administer them to the farm animal either simultaneously or sequentially.
  • the farm animals while the farm animals are fed with the suitable farm animal feedstuff ingredients, they should also be fed with the probiotic Enterococcus bacteria-containing product as described herein.
  • the farm animal product may be in form of a composition comprising the suitable farm animal feedstuff ingredients and the probiotic Enterococcus bacteria as described herein.
  • the composition is in the form of a suitable powder, a liquid or in the form of pellets or tablets.
  • the farm animal product in order to improve some stability aspects of the probiotic bacteria, it may be advantageous to provide the farm animal product as a stable emulsion of solids in-oil comprised of lipid soluble bioactive compounds such as inhibitory furanones dissolved in lipid forms of the continuous phase and with dry feed ingredients and the probiotic bacteria of interest forming the dispersed phase of the stable emulsion. See, e.g., WO 02/00035 for further details.
  • the farm animal product may be in form of a capsule, e.g., a microencapsulated product.
  • one preferred embodiment of the invention is to use Enterococcus bacteria instead ol Lactobacillus acidophilus bacteria.
  • the farm animal product comprises at least 5 times more ol Enterococcus bacteria than Lactobacillus acidophilus bacteria measured as CFU/g, in a further embodiment comprises at least 50 times more ol Enterococcus bacteria than Lactobacillus acidophilus bacteria measured as CFU/g, in another embodiment comprises at least 500 times more ol Enterococcus bacteria than Lactobacillus acidophilus bacteria measured as CFU/g and in another further embodiment comprises at least 5000 times more ol Enterococcus bacteria than Lactobacillus acidophilus bacteria measured as CFU/g.
  • one embodiment of a farm animal product does not comprise measurable amounts ol Lactobacillus acidophilus.
  • Such an embodiment of a farm animal product may be denoted a farm animal product consisting essentially of probiotic Enterococcus bacteria, or consisting essentially of at least 10 6 CFU/g of probiotic Enterococcus bacteria.
  • a farm animal product may be denoted a farm animal product consisting of probiotic Enterococcus bacteria, or consisting of at least 10 6 CFU/g of probiotic Enterococcus bacteria.
  • Another embodiment of a farm animal product does not comprise significant amounts of Lactobacillus acidophilus. In all embodiments including Enterococcus bacteria, one or more strains of such bacteria may be present.
  • the farm animal product comprises the probiotic Enterococcus bacteria in a concentration of at least 10 6 CFU/g, in a further embodiment comprises the probiotic Enterococcus bacteria in a concentration of at least 10 8 CFU/g, in a further embodiment comprises the probiotic Enterococcus bacteria in a concentration of at least 10 10 CFU/g and in a further embodiment comprises the probiotic Enterococcus bacteria in a concentration of at least 10 11 CFU/g.
  • the farm animal product comprises the probiotic Enterococcus bacteria in a concentration of less than 10 14 CFU/g.
  • the farm animal product comprises the probiotic Enterococcus bacteria in a concentration from 10 8 CFU/g to 10 12 CFU/g. In another embodiment of the invention, the farm animal product comprises the probiotic Enterococcus bacteria in a concentration of approximately 2 x 10 11 CFU/g. In a further embodiment of the invention, the farm animal product comprises the probiotic Enterococcus bacteria in a concentration of approximately 5 x 10 9 CFU/g. In a further embodiment, the farm animal product comprises probiotic Enterococcus bacteria in a concentration of approximately 5 x 10 10 CFU/g.
  • the farm animal product comprises probiotic Enterococcus bacteria in a concentration of from approximately 5 x 10 9 CFU/g to approximately 2 x 10 11 CFU/g. In another embodiment of the invention, the farm animal product comprises probiotic Enterococcus bacteria in a concentration of from 10 6 CFU/g to approximately 2 x 10 11 CFU/g.
  • the probiotic Enterococcus bacteria described herein may be any probiotic
  • the Enterococcus bacteria Based on the information disclosed herein, the skilled person is capable of selecting a specific Enterococcus strain of interest. [043] hi embodiments of the invention, the Enterococcus strain is an Enterococcus faecium.
  • Enterococcus faecium strains are SF- 273 (CHCC 4202) and SF-301 (CHCC 3828).
  • the farm animal product comprises both SF-273 (CHCC 4202) and SF-301 (CHCC 3828), and, in a further embodiment the two strains are present, in a ratio of 50:50 (based on potency).
  • the two strains are present in the farm animal product in a ratio of 60:40 or 40:60.
  • Enterococcus faecium strain number SF-273 was deposited on April 14, 1998, under ATCC 27273, in the American Type Culture Collection, 10801 University Boulevard., Manassas, Virginia 20110-2209. Also, SF-273 was deposited in the Chr Hansen Culture Collection under CHCC 4202. Enterococcus faecium strain number SF-301 was deposited on November 27, 1997, under DSM 4789, in the German Culture Collection, Deutsche Sammlung von Mikoorganismen und Zellkulturen GmbH, Mascheroder Weg lb, D-38124 Braunschweig, Germany. Also, SF-301 was deposited in the Chr Hansen Culture Collection under CHCC 3828.
  • the Enterococcus bacteria are selected to be tolerant of the following conditions: high acid (pH 4.0), high concentrations of volatile fatty acids (200 to 400 mM mixtures of acetic, propionic and butyric acids) and complete anaerobiosis.
  • Working Example 2 herein sets forth an example of one preferred assay to test if an Enterococcus bacteria is tolerant to these conditions.
  • the Enterococcus bacteria are also oxygen scavengers. Such bacteria are much more stable to oxygen exposure, moisture and heat than L. acidophilus.
  • the product bacteria are tolerant to the conditions and/or are oxygen scavengers, such traits combined are useful to confer a high degree of survival within the gastrointestinal tract of feedlot cattle.
  • the product bacteria are metabolically active upon ingestion by the animal. This activity can have an immediate impact either on the environment within the gastrointestinal tract, or on the E. coli cells present or on the receptor sites/niches with which the E. coli cells associate. Or, in another embodiment of the invention, it could be any combination of these influences.
  • the product containing the Enterococcus bacteria can be in many different forms.
  • the product containing the Enterococcus bacteria is a feed product.
  • the product containing the Enterococcus bacteria is in the form of a direct dose to the animal.
  • the product containing the Enterococcus bacteria is in the form of a direct dose to the animal through a stomach tube.
  • the farm animal may be a pig, a cow, a cattle, a sheep, a chicken, a duck or an ostrich.
  • the farm animal is a ruminant animal, in particular a cattle or a cow.
  • the farm animals are cattle.
  • the feeding may be done according to the art, and the skilled artisan is aware of how to properly feed farm animals.
  • the farm animals are fed with an amount of farm animal product that provides at least 10 7 CFU Enterococcus bacteria per animal per day, in another embodiment the farm animals are fed with an amount of farm animal product that provides at least 10 8 CFU Enterococcus bacteria per animal per day, in another embodiment the farm animals are fed with an amount of farm animal product that provides at least 10 9 CFU Enterococcus bacteria per animal per day and in another embodiment the farm animals are fed with an amount of farm animal product that provides at least 10 10 CFU Enterococcus bacteria per animal per day.
  • the farm animals are fed with an amount of farm animal product that provides less than 10 13 CFU Enterococcus bacteria per animal per day.
  • the farm animals are fed with an amount of farm animal product that provides approximately 2 x 10 11 CFU Enterococcus bacteria per animal per day. In another embodiment, the farm animals are fed with an amount of farm animal product that provides approximately 5 x 10 9 CFU Enterococcus bacteria per animal per day. i a further embodiment of the invention, the farm animals are fed with an amount of farm animal product that provides approximately 5 x 10 CFU Enterococcus bacteria per animal per day.
  • the farm animals are fed with an amount of farm animal product that provides from 10 CFU to 10 Enterococcus bacteria per animal per day, in a further embodiment the farm animals are fed with an amount of farm animal product that provides from 10 9 CFU to 10 10 Enterococcus bacteria per animal per day. In a further embodiment of the invention, the farm animals are fed with an amount of farm animal product that provides from approximately 5 x 10 9 CFU to approximately 2 x 10 11 CFU Enterococcus bacteria per animal per day.
  • the animals are fed once a day. In an alternative embodiment of the invention, they may be fed twice a day or, in another embodiment of the invention, once every second day.
  • the skilled artisan is aware of what is best in relation to a specific farm animal of interest.
  • the farm animals are fed with the farm animal product as described herein at least once a day for at least 10 days, and in a further embodiment the farm animals are fed with the farm animal product as described herein at least once a day for at least 20 days.
  • the farm animals are fed with the farm animal product as described herein until slaughter, hi a particular embodiment, the farm animals are fed with the farm animal product as described herein at least once a day for at least the last 10 days until slaughter, and in another embodiment the farm animals are fed with the farm animal product as described herein at least once a day for at least the last 20 days until slaughter.
  • the farm animals are fed with the farm animal product as described herein for about 26 days.
  • the farm animals are fed for about 26 days with an amount of farm animal product that provides from approximately 5 x 10 9 CFU to approximately 2 x 10 11 CFU Enterococcus bacteria per animal per day.
  • the farm animals are fed for about 26 days with an amount of farm animal product that provides from approximately 5 x 10 9 CFU to approximately 5 x 10 10 CFU Enterococcus bacteria per animal per day.
  • the farm animals are fed for about 26 days prior to slaughter with an amount of farm animal product that provides from approximately 5 x 10 9 CFU to approximately 5 x 10 10 CFU Enterococcus bacteria per animal per day. Additionally, in this embodiment, the animals can be fed the farm animal product set forth in Example 3.
  • the farm animal product is used for feeding the animals in an amount and for a number of days where the farm animal product reduces the number ol Escherichia coli O157:H7 cells quantified in the faeces of the challenged animals by at least 1.5 logs, or, in another embodiment, by at least 2 logs.
  • the farm animal product reduces the number ol Escherichia coli O157:H7 cells quantified in the faeces of the challenged animals by at least 1.5 logs in the first few days of treatment, or, in another embodiment, by at least 2 logs in the first few days of treatment, hi another embodiment of the present invention, the farm animal product reduces the number ol Escherichia coli O157:H7 cells quantified in the faeces of the challenged animals by at least 1.5 logs in 3 days of treatment, or, in another embodiment, by at least 2 logs in 3 days of treatment.
  • the farm animal product reduces the number ol Escherichia coli O157:H7 cells quantified in the faeces of the challenged animals by at least 1.5 logs in 10 to 14 days of treatment, or, in another embodiment, by at least 2 logs in 10 to 14 days of treatment. In yet another embodiment of the present invention, the farm animal product reduces the number ol Escherichia coli O157:H7 cells quantified in the faeces of the challenged animals by about 1.5 logs to about 2 logs.
  • the farm animal product reduces the number ol Escherichia coli O157:H7 cells quantified in the faeces of the challenged animals by about 1.5 logs to about 2 logs within 3 days of treatment. Additionally, in this embodiment, the animals can be fed the farm animal product set forth in Example 3.
  • the product containing the Enterococcus bacteria can be administered to the animal in many ways.
  • the product containing the Enterococcus bacteria is fed to the animal as a feed product.
  • the product containing the Enterococcus bacteria is dosed directly to the animal.
  • the product containing the Enterococcus bacteria is dosed directly to the animal through a stomach tube.
  • Example 1 Dose Titration Study to determine the effect of two different dosing regimens of a characterized Direct Fed Microbial Culture on the post-challenge fecal shedding of Escherichia coli 0157: H7.
  • CHB DFM Two Enterococcus faecium strains present together at 50:50 (based on potency) in a gelatin bolus.
  • the strains are SF-273 (CHCC 4202) and SF-301 (CHCC 3828).
  • the two Enterococcus faecium strains each are present in the product at 2.5 x 10 9 CFU/g.
  • CHB Probios TC A corresponding product, which comprises probiotic Enterococcus faecium bacteria in a similar CFU/g as for CHB DFM. This product also comprised active dry yeast at around 2.5 x 10 9 CFU/g.
  • Treatment D Probios TC at a rate of 2 grams/head/day • Treatment ⁇ : Probios TC at a rate of 20 grams/head/day
  • coli O157:H7 (5.2 x 10 10 CFU/animal). Following administration of the challenge material, the tube was flushed with 120 mL of sterile phosphate buffered saline. [072] Animals were evaluated daily for evidence of adverse reactions. Fecal (rectal) specimens were collected on 1, 3, 5, 8, 10, 12, 14, 17, 19, 23, 24, 26, and 30 days following oral challenge. Fecal samples were placed in whirl packs, packed in ice and provided for detection and quantification of Nal r E. coli 0157: H7.
  • HE. coli O157:H7 colonies were not detected by direct plating (detection limit > 10 2 /g), GNccv broth incubated for 18 to 24 h was plated, in duplicate, on SMAC plates containing Nal (20 ⁇ g/ml) and incubated for 24 h at 37°C. Following incubation, three colonies per sample with typical colony morphology (from the enriched samples) were streaked on blood agar plates and incubated for 24 h at 37°C. The indole test was done on colonies from the blood agar plates and indole positive colonies were tested for agglutination specific for 0157.
  • the independent variables were treatment, day, and the treatment by day interaction.
  • the comparison was done using repeated measures analysis of variance (MIXED procedure, SAS Institute Inc.).
  • the outcome was the level of fecal shedding (CFU/g of faeces) with group as the treatment and day as the repeated measure. Day and treatment interaction was included as a fixed effect. Colony counts were log transformed prior to analysis. Results:
  • Treatment by Days Interaction P 0.6 a, b Means not sharing the same letters differ at P ⁇ 0.1.
  • Example 2 Assay to select Enterococcus bacteria that are tolerant to preferred conditions.
  • the medium of Bryant MP and Burkey LA (J. Dairy Science 1960) containing 40% rumen fluid (RF medium) was prepared either under 100% CO2 (with sodium carbonate solution as buffer) or under 80%:20% N2:CO2 headspace gas (with sodium bicarbonate solution as buffer). This medium was used to score growth of the test strains as 0 (no growth) or +, ++, +++, or ++++ (excellent growth) at 37C after 48 to 72 h.
  • the inoculated tubes were exposed to the atmosphere for 5 min then closed and incubated overnight (ca. 18 h) at 37C. After incubation the top one-centimeter of the medium had oxidized (turned the redox indicator pink) and the resultant growth pattern of each test strain was scored. The strain was scored as a strict anaerobe if visible growth along the stab line was only in the reduced portion of the tube. The strain was scored as an oxygen tolerant anaerobe if strain growth extended into the pink (oxidized) portion of the medium. The strain was scored as a facultative anaerobe if growth was visible along the stab line within both the reduced and oxidized portions of the tube.
  • the strain was scored as an aerobe if growth was visible only within the oxidized portion of the medium.
  • an Enterococcus bacterium is considered tolerant to complete anaerobiosis conditions if it grew in the reduced portion of the medium.
  • Example 3 Formulation of farm animal product.
  • Table 4 sets forth the composition of one embodiment of a farm animal product according to the present invention, hi this embodiment depicted in Table 4, the farm animal product is not less than 200 billion CFU/gram.
  • the farm animal product in Table 4 is packaged in 250 gm foil pouches, with 14 pouches per case, and it is available from Chr. Hansen, Inc. in Milwaukee, Wisconsin under the name Probios FS.
  • Table 4 Farm animal product containing probiotic Enterococcus bacteria.
  • Dried Enterococcus faecium Fennentation Product (45.0-75.0%) (50:50 mix of E. faecium strains SF-273 and SF-301)

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Abstract

L'invention concerne un produit d'animaux d'élevage contenant une bactérie probiotique Enterococcus et l'utilisation de ce produit pour diminuer le nombre de cellules pathogènes Escherichia coli O157:H7 chez les animaux d'élevage tels que les bovins.
PCT/DK2004/000519 2003-07-30 2004-07-30 Produit d'animaux d'elevage contenant une bacterie probiotique enterococcus WO2005009139A1 (fr)

Applications Claiming Priority (6)

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US49094303P 2003-07-30 2003-07-30
US60/490,943 2003-07-30
US49756403P 2003-08-26 2003-08-26
EP03077658.7 2003-08-26
EP03077658A EP1510135A1 (fr) 2003-08-26 2003-08-26 Aliment pour animaux de fermes comprenant enterococci probiotiques
US60/497,564 2003-08-26

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1989011858A1 (fr) * 1988-06-09 1989-12-14 Whitecliffe Laboratories Limited Traitement de l'immunodeficience
WO1993006208A1 (fr) * 1991-09-20 1993-04-01 Pioneer Hi-Bred International, Inc. Enterococcus microencapsule dans un acide gras destine a la volaille
EP0856259A1 (fr) * 1996-12-23 1998-08-05 SITIA-YOMO S.p.A. Composition pour l'usage dans l'alimentation comprenant de bactéries vivantes lactiques lyophisées
US6503544B2 (en) * 2000-08-21 2003-01-07 Nisshin Feed Inc. Animal feed additives

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
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