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    • 专利摘要:
    The invention relates to the use of at least one bacterium strain of the genus Bacillus and at least one lactic acid bacterium for the prevention or treatment of pathogens affecting the health of farm animals.
    公开号:FR3073139A1
    申请号:FR1760495
    申请日:2017-11-08
    公开日:2019-05-10
    发明作者:Michel Layus；Alexandre Brame；Stephane Thebault
    申请人:Nolivade SARL；
    IPC主号:
    专利说明:

    Composition improving animal welfare
    The invention relates to a composition for improving the welfare of animals, in particular farm animals.
    Health problems in farms are often the consequence of the combination of several pathogens. These can be present in different microbial ecosystems: the environment (air, organic or inorganic surfaces), in the internal system of the animal (digestive or respiratory system) or on the animal (cutaneous system).
    In the different types of land animal breeding, there are always risky phases where microbial ecosystems evolve strongly, such as for example a phase of re-implantation of an external flora after disinfection, the arrival of animals in a farm , the first moments of life of newborns, the arrival of new individuals in a batch of already existing animals, food transitions, the phase of reimplantation of an internal flora after use of antibiotics etc ...
    Also, it is necessary to control this environment in order to maintain the sanitary condition of the farms and preserve the welfare of the animals.
    Antibiotic treatment strategies have been used for a very long time but pose two major problems: on the one hand, the resistance generated by the too regular use of antibiotics, and on the other hand, a public health problem by the consumption of chairs animals containing many antibiotics.
    Also, there is a need to find alternatives to antibiotics.
    A more recent approach has been developed aiming to use bacterial flora making it possible to regulate the pathogenic flora found in farms. Mention may be made, for example, of applications in the environment as in application EP0852114, where non-pathogenic bacteria are used to propose a systematic disinfection process for buildings intended for raising a herd.
    Other approaches are used and aim to directly treat animals by the dermal route with non-pathogenic bacteria. Mention may in particular be made of application CN104307012 which proposes deodorant and sanitizing compositions which can be sprayed on poultry, or else application CN 103773721 which proposes probiotic solutions limiting the risks of diarrhea in newborns, the examples of application being for example a spray on the mother's udders.
    However, despite these different methods, the health results do not seem satisfactory.
    Also, there is a need to find a solution to maintain the breeding environment of healthy terrestrial animals, in order to promote their health and thus their well-being, which will promote the productivity of farm animals.
    The purpose of the invention is to overcome the above-mentioned drawbacks.
    Another object of the invention is to provide a new, effective method for maintaining the health and well-being of farm animals.
    Another object of the invention is to provide a kit which can be used directly by breeders.
    Also, the invention relates to a bacterial composition comprising at least one strain of bacteria of the genus Bacillus, in particular of Bacillus subtilis and at least one strain of lactic bacteria, in particular at least one strain of Lactococcus lactis, for its use in the context of prevention or treatment of pathogenic infections affecting the health of terrestrial farm animals, said composition being:
    at. Ingested or inhaled by said land animals, and
    b. is
    b.1. spread in the breeding environment of said terrestrial farm animals,
    b.2. applied to said land animals,
    b.3. applied in the culture medium of said terrestrial farm animals and applied to said terrestrial farm animals.
    The invention is based on the surprising observation made by the inventors that a mixture of at least one strain of bacteria of the genus Bacillus subtilis and at least one strain of the genus Lactococcus lactis, non-pathogenic bacteria, act synergistically on the animal health of terrestrial animals when the said mixture is both ingested or inhaled by the animals and either spread in the breeding environment, or applied directly to the animal, or both. In the invention, the spreading is assimilated to a dispersion; these two terms can be used uniformly in the invention.
    The inventors have proven a synergy of the protective effect against pathogens when the bacteria described in the invention (Bacillus and Lactococcus) are both present in the digestive or respiratory tracts and in the breeding environment and / or on the 'animal.
    In the invention, the term “bacterial strain” is understood to mean a group of individuals resulting from successive subcultures of a bacterial colony. A bacterial strain is a part of a bacterial species different from other bacteria of the same species by a minor but identifiable difference.
    In the invention, by pathogenic infections is meant infections by the following microorganisms: Salmonella, Staphylococci, Streptococci, Clostridium, Listeria, Arcanobacterium, Mycoplasma, Klebsielle, Pasteurelles, Bordetelles, Campylobacter, Brachyspire, Escherichia coli and / or responsible for the pathologies following: enteritis, salmonellosis, ulcers, lameness, rhinitis, mastitis, epidermitis, necrosis, pasteurellosis, bordetellosis, listeriosis.
    It is particularly advantageous in the invention that the above-mentioned composition comprises at least two strains of Bacillus, and in particular at least two different strains of Bacillus and at least one strain of lactic acid bacteria, in particular of Lactococcus lactis.
    It is even more advantageous in the invention for the above-mentioned composition to comprise at least three strains of Bacillus, and in particular at least three different strains of Bacillus, and at least one strain of lactic acid bacteria, in particular of Lactococcus lactis.
    It is even more advantageous in the invention that the above-mentioned composition comprises at least four strains of Bacillus, and in particular at least four different strains of Bacillus and at least one strain of lactic acid bacteria, in particular of Lactococcus lactis.
    In the invention, an application in the environment (E), with reference to spreading in the environment, will be distinguished from an external application on the animal (A) with reference to the application on the animal, and an internal application (I), with reference to ingestion or inhalation.
    Application E makes it possible to control and / or orient the bacterial flora of the environment of farm animals by reducing the pathogenic pressure. It can be done by nebulization or spraying of livestock buildings, incorporation into the litter ... We observe less contamination of animals because there are fewer pathogenic foci. Such an application plays on living or surviving pathogens in the environment. The methods of spreading in the environment are spraying, nebulizing, covering surfaces with foam or even dusting with powder.
    The spreading is generally done on the surfaces of the farm, mainly on surfaces in direct or potential contact with animals, preferably on cleaned and disinfected surfaces, after having waited for the end of the persistence of disinfectant products.
    Land application in the environment is carried out at least once after a crawl space, after a cleaning / disinfection protocol or before the animals arrive. It is particularly advantageous to make a repeated application every week.
    Application A makes it possible to control or orient the microbism of the animal by reducing the pathogenic pressure. The application can be made by nebulization, spraying, soaking of the terrestrial animal. This generates less contamination of animals because there are fewer pathogenic foci directly on the animal.
    The application on the animal can be done by spraying, nebulization, soaking, footbath, foam, depending on the area of the animal on which it is desired to deposit the composition according to the invention.
    It is advantageous to apply the composition at least once to the animal as early as possible in the life of the animal or when arriving in a new environment. Preferably the application is repeated every week.
    Application I makes it possible to control and / or orient the digestive microbism by reducing the pathogenic pressure and / or by stimulating the animals' natural defenses. The application can be done via drinking water, food or any means allowing the animal to ingest the composition of the invention. In this way, the composition according to the invention plays directly on the digestive health of land-based animals, which limits the entry of other pathogens.
    Application I can also be an application by inhalation by nebulization or spraying into the respiratory tract.
    Application I should be started as soon as possible, before the start of the health risk phase and for the duration of the health risk period. It is recommended to renew the application at least every 10 days, at best every day.
    In the invention, the term “farmed animals” means all the animals which are not aquatic, and which can be used directly or indirectly for human or animal nutrition. This concerns in particular cattle, sheep, pigs, goats, rabbits, equines, insects but also farm birds such as chickens, ducks, geese, turkeys, quail and other gallinaceous animals. 'breeding.
    Advantageously, the invention relates to a composition for its use mentioned above, in which the bacterial composition comprises:
    - at least one of the following three Bacillus subtilis strains: NOL01, NOL02, NOL03, said strains being respectively deposited at the CNCM under the number CNCM I - 4606, CNCM I-5043 and CNCM I - 4607, and
    - at least the strain of lactic bacteria: Lactococcus lactis spp lactis 1 strain NOL11, said strain being deposited at the CNCM under the number CNCM I - 4609.
    The invention relates to the use of at least one strain chosen from the NOL01, NOL02 and NOL03 strains and at least the NOL11 strain. Also, the invention covers the following 7 combinations:
    - NOL01 and NOL11,
    - NOL02 and NOL11,
    - NOL03 and NOL11,
    - NOL01, NOL02 and NOL11,
    - NOL01, NOL03andNOL11,
    - NOL02, NOL03andNOL11, and
    - NOL01, NOL02, NOL03 and NOL11.
    All these strains were deposited in the national collection of microorganisms (CNCM) at the Pasteur Institute in Paris, according to the Budapest treaty.
    More advantageously, the invention relates to a composition for its use above, said composition comprising from 10 4 to 10 11 bacterial colonies of Bacillus and from 10 4 to 10 11 bacterial colonies of lactic acid bacteria, the bacterial colonies being in grams or in milliliter of composition.
    In other words, in this advantageous embodiment, if the composition according to the invention is in liquid form, said composition will comprise from 10 4 to 10 11 bacterial colonies of Bacillus per ml of composition, and this for each of the strains when the composition comprises at least two strains and from 10 4 to 10 11 bacterial colonies of Bacillus per ml of composition of lactic bacteria.
    If, on the other hand, the composition according to the invention is dehydrated or nonaqueous, said composition will comprise from 10 4 to 10 11 bacterial colonies of Bacillus per g of composition, and this for each of the strains when the composition comprises at least two strains and from 10 4 to 10 11 bacterial colonies of Bacillus per g of lactic acid bacteria composition.
    In the invention, from 10 4 to 10 11 bacterial colonies means: approximately 10 4 , approximately 5. 10 4 , approximately 10 5 , approximately 5.10 5 , approximately 10 6 , approximately 5.10 e , approximately 10 7 , approximately 5.10 7 , approximately 10 8 , approximately 5.10 8 , approximately 10 9 , approximately 5.10 9 , approximately 10 10 , approximately 5.10 10 or approximately 10 11 bacterial colonies.
    Bacterial colonies are defined as mL (or gram) of bacterial culture. A person skilled in the art easily knows how to determine this number of bacteria, in particular by counting either manually (using a Malassez slide) or by using an automatic cell counter, or by dilution and then seeding on agar and counting of colonies.
    Advantageously, 10 5 to 10 10 bacterial colonies will be used per individual when treating poultry, 10 5 to 10 10 bacterial colonies per individual for application in piglets, and 10 5 to 10 11 bacterial colonies per individual for application in pigs.
    Regarding the advantageous dose for ingestion, this is 10 5 to 10 10 bacterial colonies per animal per application.
    For the environment, it is advantageous to use 10 5 to 10 11 bacterial colonies per m 2 of surface or environment to be treated.
    Even more advantageously, the invention relates to a composition for its use as defined above, wherein said at least one strain of Bacillus is in sporulated form and / or in vegetative form.
    Lactic acid bacteria are always in vegetative form. Also the composition according to the invention comprises at least one strain of Bacillus in sporulated form and at least one strain of lactic acid bacteria in vegetative form, or comprises at least one strain of Bacillus in vegetative form and at least one strain of lactic bacteria in form vegetative.
    Even more advantageously, the invention relates to a composition for its use defined above, wherein said composition inhaled or ingested and said composition spread in the breeding medium of said terrestrial farm animals and / or applied to said farm animals terrestrial, are used simultaneously, separately or spread over time.
    Treatments that are ingested or inhaled and treatments that are applied to the environment or applied to animals do not have to be simultaneous, that is, applied at the same time. It is possible to separate the applications over time, for example to start treating the animal's environment, then once the animal has been introduced into the environment, have it ingest the composition according to the invention.
    More advantageously, the invention relates to a composition for its aforementioned use, wherein said composition is spread and / or applied prior to ingestion or inhalation by said land animals.
    In the case of a treatment prior to ingestion or inhalation of the composition according to the invention, spreading in the environment is carried out according to the doses as defined above.
    Advantageously, the invention relates to a composition for its use mentioned above, in which said composition consists essentially of the four bacterial strains of the group consisting of the three strains NOL01, NOL02, NOL03, said strains being respectively deposited at the CNCM under the number CNCM I 4606, CNCM I-5043 and CNCM I - 4607 and the strain Lactococcus lactis spp lactis 1 strain NOL11, said strain being deposited at the CNCM under the number CNCM I - 4609.
    This combination is particularly advantageous and described in the examples.
    It is particularly advantageous that the composition comprises the three strains of Bacillus subtilis bacteria mentioned above, in vegetative or sporulated form, and the lactic acid strain mentioned above, in vegetative form.
    Even more advantageously, the invention relates to a composition for its use as defined above, wherein said composition is spread by m 2 at the rate of 10 5 to 10 11 bacterial colonies.
    In the context of the application of the environment of farm animals, it is advantageous for the composition to be dispersed or spread at a rate of 10 5 to 10 11 bacterial colonies for a dry or dehydrated composition, or 10 5 to 10 11 bacterial colonies for a liquid composition per m 2 of surface or medium to be treated.
    Still advantageously, the invention relates to a composition for its aforementioned use, in which the environment is cleaned and disinfected and the terrestrial animals are washed before spreading in the breeding medium of said terrestrial breeding animals and / or the application to said terrestrial farm animals.
    It may be advantageous to clean the environment and / or the animals before using the composition according to the invention. This cleaning can be considered as disinfection. Before implantation of the flora of the composition according to the invention, if cleaning or disinfection is carried out, the better the effectiveness of the bacteria, and the better the benefit for the animals. The surfaces can be treated by any cleaning means known to a person skilled in the art using the composition according to the invention. As regards the cleaning of animals, it is possible to wash them, bathe them, or even shampoo them with solutions comprising the composition according to the invention.
    The invention further relates to a kit for terrestrial farm animals against pathogenic infections affecting the health and well-being of farm animals comprising
    - a composition as defined in any one of the preceding claims, and
    - at least one biosecurity agent and / or at least one biocontrol agent
    In the invention, the term biosecurity agent means a chemical or physical agent capable of reducing the number or inactivating microorganisms and their biofilms. These include disinfectants, detergents, UV radiation, or even steam or any other biocidal agent. This list is not exhaustive, and the skilled person knows the biosafety officers.
    In the invention, the term “biocontrol agent” means any composition of living organisms or derived from living organisms, or composition of natural substances whose purpose is to protect animals from a health point of view (macro-organisms (insects, nematodes, etc.), microorganisms (viruses, bacteria, phages or fungi), chemical mediators (pheromones), or natural substances of mineral, vegetable or animal origin such as for example essential oils).
    In the invention, it is proposed to provide a kit which comprises the composition as defined above, and in particular a composition comprising the bacteria NOL01, NOL02, and NOL03 with the bacteria NOL11, and in parallel at least one biocontrol agent and / or biosecurity.
    This kit allows the user to have the means to apply the composition according to the invention, and if necessary, before or after treatment with the composition according to the invention, to use one or more other agents such as defined above.
    It is notable that to maintain the effect of the composition according to the invention, the abovementioned agent should not be used simultaneously with the composition according to the invention if it has an antibacterial effect.
    The invention will be better understood in the light of the three examples and the following 13 figures:
    BRIEF DESCRIPTION OF THE FIGURES
    Figure 1 represents a graph representing the results of corrected consumption index (ICC) of the test and control livestock buildings. The abscissa axis differentiates the results of the test lots (left) and the control lots (right). The y-axis represents the ICC values. For all of the test lots, identified by the black dots in the left column, a synthetic mustache box is represented to visualize means, medians, quantiles and confidence indices. Likewise for the control batches in the left column. A .: invention; B: witnesses.
    Figure 2 represents a graph representing the difference in corrected consumption index (CCI) results between the test and control buildings on the same farm. The y-axis represents the ICC values. For all the farms, identified by the black dots, a synthetic mustache box is represented to visualize averages, medians, quantiles and confidence indices.
    Figure 3 represents a graph representing the results of the individual weighings of animals at 7 days of age, all farms combined, in the test and control buildings. The abscissa axis differentiates the results of the test batches (on the left -A.) And the control batches (on the right - B.). The ordinate axis represents the values of the weights in kg. A synthetic mustache box indicates the values of the means, medians, quantiles and confidence indices for all of the animals from the trial (left) or control (right) lots.
    Figure 4 represents a histogram representing the results of burnt tarsi obtained at the slaughterhouse for female hens. The abscissa differentiates the control lots (light gray on the left) and tests (dark gray on the right). The ordinate represents the percentage of tarsi burned. The graphs show the value of the medians of the two lots and their confidence interval.
    Figure 5 represents a histogram representing the results of burnt tarsi obtained at the slaughterhouse for male chickens. The abscissa differentiates the control lots (light gray on the left) and tests (dark gray on the right). The ordinate represents the percentage of tarsi burned. The graphs show the value of the medians of the two lots and their confidence interval.
    Figure 6 represents a histogram representing the results of consumption of antibiotic in ppm of antibiotic per kg of bodyweight. The abscissa differentiates the control lots (light gray on the left) and tests (dark gray on the right). The ordinate represents the antibiotic consumption value. The graphs show the value of the medians for the two lots and their confidence interval.
    Figure 7 represents a histogram representing the number of days of antibiotic treatment accumulated over all of the test or control lots. The abscissa differentiates the treatments carried out for problems related to lameness (A.) or the treatments carried out in the first 10 days of life of the animals (start B.). The ordinate represents the number of days of antibiotic treatment. The results for the test lots are distinguished in light gray and the control lots in dark gray.
    Figure 8 shows a histogram showing the average weight results obtained at the slaughterhouse for male chickens. The abscissa differentiates the control lots (light gray on the left) and tests (dark gray on the right). The ordinate represents the average weight of the animals. The graphs show the value of the medians of the two lots and their confidence interval.
    Figure 9 represents a histogram representing the results of average weights obtained at the slaughterhouse for female hens. The abscissa differentiates the control lots (light gray on the left) and tests (dark gray on the right). The ordinate represents the average weight of the animals. The graphs show the value of the medians of the two lots and their confidence interval.
    Figure 10 represents the modeling of the evolution of the cumulative mortality in female hens. The abscissa represents the days of life of the animals and the ordinate the percentage of cumulative deaths. In light gray (above) the model corresponding to the control batches is displayed and in dark gray (below) the model of the test batches.
    Figure 11 represents the modeling of the risk of occurrence of diarrhea in a cell according to the protocol followed. The abscissa axis represents the days of life of the piglets compared to the birth date (0). The ordinate represents the probability that at least one piglet in a pen has diarrhea. The three grouped curves located above represent the models of the diarrhea probabilities for lots 1 to 3 and the isolated curve below for lot 4.
    Figure 12 represents the modeling of the risk of mortality of a piglet according to the protocol followed. The abscissa axis represents the days of life of the piglets compared to the birth date (0). The ordinate represents the probability that a piglet dies. The three grouped curves located above represent the models of the diarrhea probabilities for lots 1 to 3 and the isolated curve below for lot 4.
    Figure 13 represents a graph representing the number of piglets with normal faeces out of 120 boxes with shampoo and 120 boxes without shampoo. The abscissa represents the days of life of the piglets and the ordinate the number of boxes with normal faeces. In light gray, we visualize the boxes that did not receive shampoo before receiving the flora and in black the boxes that received the shampoo before receiving the flora.
    EXAMPLES
    EXAMPLE 1 - application in the environment: litter from farmed calves
    In this first example, the inventors seek to show whether an application of the composition according to the invention in the environment or on the litter alone is of interest for the prevention of pathogenic infections.
    This test is carried out on the calves part of a Holstein dairy cow farm in France in the department of Ille-et-Vilaine.
    The product tested is an association of living bacteria (strains of Bacillus subtilis and Lactococcus lactis) in their culture medium, brought frozen to the farm, to be diluted in water before spraying.
    The test is carried out in the calf building with a control batch and a batch where the product will be sprayed into the environment and onto the litter. The impact of the product will be studied over time.
    The influence of the product will be measured on the chemical qualities of the litter, the cleanliness of the animals and the health balance of the batch (frequency and nature of the pathologies, veterinary costs) over time.
    MATERIALS AND METHODS a - Holstein calf animals from 6 weeks to 5 and a half months divided by 3, 4 or 5 according to their age in 4 huts of 15m 2 in straw area. The first two boxes are separated from the other two by a wall.
    b -Product used and material
    A dose of composition is packaged in two separate 20 ml vials:
    • Bottle B containing bacillus • Bottle L containing Lactococcus
    The bottles are brought frozen in breeding.
    The water used during spraying must not contain chlorine, a preliminary treatment of water with sodium thiosulfate may therefore be necessary.
    The equipment required for processing and measurements is as follows:
    - 8 boxes of 2 composition vials
    - Sodium thiosulfate
    - 1 sprayer
    - 1 bovimetric tape c - Experimental device
    BEFORE LAUNCHING THE TESTS:
    - Take care to check that the sprayer is working properly.
    - Total surface of the mulched area (control + test): 60 m 2 , or approximately 4 m 2 per animal
    - Surface treated with the composition according to the invention: 2 boxes of 15 m 2
    - Control surface: 2 boxes of 15 m 2
    Separation of control boxes and tests by a wall.
    The weight of the calves is estimated at the start of the test using a bovimetric tape. The calves were distributed in each box to obtain a total live weight comparable between the control and test boxes.
    d - Conduct of the test and measurements
    The trial lasted 2 months, a period between two treatments.
    Curing Spraying of boxes 3 and 4
    Sem Sem Sem Sem Sem Sem Sem Sem
    17 18 19 20 21 22 23
    Sem
    X X X X X X X X X X
    The first application of the product is made after cleaning in week 16. The first measurements are made just before the first application of the flora. Then the measurements are carried out every week before applying the product. End of product applications in week 23 and last measurement just before the second cleaning in week 24.
    e -Application
    Directions for use: apply in liquid form. A dose of product (consisting of bottles B and L) is necessary for each weekly spraying.
    L of diluted composition is used to treat the 30 m 2 of the calf test.
    Solution preparation procedure:
    • Let the product thaw at room temperature between 15 and 25 ° C until complete liquefaction (approx. 1h ± 15 minutes) • Prepare 10L of water and add 160mg of sodium thiosulfate (if the water is treated with chlorine, add sodium thiosulfate up to 16 mg / L of water (if the water is treated with hydrogen peroxide, it is essential to use mineral water or untreated water).
    • Remove the cap and remove the stopper from the vials • Pour the two vials of the product into the treated water • Mix to homogenize the product then leave to stand for 15 minutes • In a sprayer that did not contain any disinfectant, pour 1L of the composition according to the invention, said composition being diluted, and spray on the litter and all the surfaces of boxes 3 and 4.
    • This application will be renewed every week of the trial (8 in total).
    During the test, no disinfection will be carried out in the building.
    f- Mulching practices
    The farmer keeps his usual mulching practices throughout the duration of the test.
    • Bring 1 round of straw for 30 m 2 after cleaning. • Mulching 2 to 3 times a week • Distribute the quantities of straw identically in each of the boxes.
    • During measurement / treatment days, the practices g - Measurements and Analyzes
    Date of samples and measurements:
    mulching is carried out after these
    Sem Sem Sem
    17 18 x
    Sem
    Sem
    Sem
    Sem
    Sem
    Week x
    flushing
    MS of litter T ° of litter Height of litter
    XXXXX X X X X X X X X X X X X X X X X X X
    • Temperature measurements
    Take a weekly T ° measurement on the horizon 0 - 20 cm from the first cleaning on the surface of the straw and in depth (indicate the depth of the first measurement to make the same throughout the test, if the 'thickness increases, a third deeper measurement can be taken (total length of the probe).
    • MS measurements
    From the first cleaning, once every two weeks, take litter samples (3-4 handles for a total of 200g minimum) on the horizon 0-20 cm, and measure the DM (sending samples to the laboratory) . In the weeks with cleaning, perform the measurement on the day of cleaning, just before the start of operations.
    For each sample, take a sample at all the sampling points, and collect the samples by box. We thus obtain 4 samples of 1 kg minimum. Homogenize before sending for analysis and identify as follows: box number (1,2,3 and 4) / date of sampling / analysis (MS) • Measurements on animals
    Sanitary: ask the farmer to note the sanitary problems which appeared during the test + the veterinary costs for each period (diarrhea, respiratory problem or other).
    Results and conclusion:
    The results of the litter temperatures as well as the dry matter are equivalent between the two batches.
    Some diarrhea observed, but no significant difference between the control batch and test batch (treated with the composition according to the invention), in particular in terms of faeces and in terms of cleanliness of the huts where the results are equivalent between the batches.
    Also, application in the environment alone is not sufficient to improve the living conditions of animals.
    EXAMPLE 2 - Application to animals - poultry
    BACKGROUND AND OBJECTIVES
    In this second example the objective is to determine if the application on the animal can have a better effect. The objective of this trial is to test the application of barrier flora (frozen combination of Bacillus and Lactococcus) from the hatchery, on OJ chicks, in order to study its impact on the sanitary quality of batches. The test will be carried out on chicks destined for 20 flocks, which will be compared to contemporary control batches. In each farm under test, only the chicks from one building will be treated in a hatchery, the chicks from the second building will not have undergone any treatment (control).
    The goal is to verify an effect on the decrease in mortality, health improvement, improvement of zootechnical indices.
    The composition according to the invention (strains of Bacillus subtilis and Lactococcus lactis) is applied directly to the chicks after sexing. The doses will be thawed in a water bath of 27 ° C for 9 minutes 30.
    They will then be diluted in 3L of mineral water for 30,000 chicks, or 0.1 mL per chick. This recommendation takes into account the volume of water used for a spray vaccination, ie 0.2 to 0.3mL.
    100 males and 100 females are weighed on D7 in each building.
    The parameters studied are: food and water consumption, mortality and elimination as well as data from the slaughterhouse (technical and economic results, seizures, counting of pododermatitis, tarsi, LCI and brechets).
    Results:
    The results observed for the corrected consumption index (CCI) are shown in Figures 1 and 2.
    With a Student test: p-value = 0.882, the difference between the treated and untreated animals is not significant.
    It is the same for the feed chick margin (MPA), the burnt tarsi, the mortality, the antibiotic treatments, the weights of the animals mentioned in FIG. 3, the pododermatitis which are not significantly different between the controls and the animals having received the composition according to the invention.
    Also, we see that the application on the animal alone does not bring any benefit to the animal.
    EXAMPLE 3 - Application in the environment, on animals: Poultry
    GOAL
    The objective of this test is to test the effectiveness of the composition according to the invention both on animals and in the environment. The aim is to identify an optimal use for the prevention of pathologies (lameness, colibacillosis, etc.) and / or intestinal health and the improvement of the quality of litter and therefore animal welfare.
    EXPERIMENTAL DEVICE tested farms.
    The first farm tested has 2 comparable buildings, with buckwheat husk litter on concrete floor. It is equipped with scales. Its drinking water is treated with 3% 80% pure hypochlorous acid (electrolysis treatment system - company DCW).
    The second farm has 2 comparable buildings, with buckwheat husk litter and wood chips on non-concrete soil. It is equipped with scales. Its drinking water is treated with hydrogen peroxide, stopping the treatment when the flora is sprayed.
    The influence of the product is measured on growth performance, the health balance of the lot (frequency and nature of pathologies, veterinary costs), bacteriological counts (litter), gas emissions, quality of litter and good criteria -to be.
    APPLICATION
    Application of 5 doses during breeding.
    doses distributed as follows: D0-D3 J7-J9-J14-D21.
    The seeding on D0 is carried out after the establishment.
    Spray the environment and animals.
    The test and control buildings are distinguished only by the application or not of the barrier flora. The product is sprayed by misting (approximately 15-20L of water containing the flora for each application, for 1200 m 2 ). The water used is untreated artesian well water.
    Results:
    The following table presents a comparison of the results between the control and test buildings for the two farms.
    Breeding 1 Breeding 2 Zootechnical performances ICC +4 pts +2.5 pts Welfare Water / Food + before 28d- after 28 daysLitter - before 28d - at the end of the tape Dry matter - except at the end of the tape - laminitis - + Sanitary state treatments more treatments in the control batch several treatments in both batches Mortality + for males + for females before 27 d then -- for males after 27 days Sorting + for males - Held on legs + - seizures + for females - Bactérioloqie Litter No significant difference No significant difference
    Conclusions:
    Animal and environmental applications alone are not enough to improve animal welfare. Indeed, the results on the 2 farms show contradictory effects on lameness, mortality and pododermatitis, no significant difference on bacteriology and litter and a degraded consumption index.
    EXAMPLE 4 - Applications in the environment, on animals and ingested: Poultry
    The purpose of this trial is to determine whether a combination of environmental, animal and ingestion applications is effective in preventing pathogenic infections.
    at. Experimental apparatus
    A preliminary statistical study made it possible to classify broiler poultry farms according to the proximity of the zootechnical results of two livestock buildings. Have entered this classification, all farms with at least 5 bands since the beginning of 2014 for which the values of weight, mortality, CHF and pododermatitis have been entered. This study made it possible to establish a list of the 60 farms with the closest results between two buildings.
    The test will therefore take place in around twenty farms on this list, with each control farm, a control building and a building receiving an application of barrier flora. Before integrating each farm into the test, it will be ensured that the two buildings under test will receive the same lines from the same hatchery for 2 strips.
    b. Product used and material
    The product will be applied with rechargeable sprayers or with farmers' sprayers used for vaccination (after validation of the equipment: sufficient capacity, practicality, no use for disinfection treatments or any other incompatible product).
    The equipment required for processing and measurements is as follows:
    Boxes of 2 bottles of flora
    Mineral water
    Accumulator nebulizer
    Gloves
    On Boot
    Cottes
    vs. applications
    Two types of application are carried out: in the building + animals and in the water circuit.
    Environment and animals application:
    Spraying on: litter, feeders, drinkers and animals.
    dose for 1000 to 1200 m 2 or 1 dose for 20 to 24,000 chicks.
    Application: D0 (just after installation), D10 and D20.
    Solution preparation procedure:
    Leave the product to thaw at room temperature between 15 and 25 ° C until complete liquefaction (approx. 1h ± 15 minutes) or 5 minutes in a 30 ° C water bath.
    Pour 10L of water in the spray tank (if the water is treated with chlorine, add sodium thiosulfate up to 16 mg / L of water. If the water is treated with hydrogen peroxide, use imperatively mineral water or untreated water).
    Remove the cap and remove the stopper from the vials
    Pour the two bottles of the product into water (treated or mineral)
    Mix to homogenize the product then let stand for 15 minutes before starting the nebulization. Spray the solution throughout the building and on animals.
    Application in the water circuit:
    Water treatment stopped (or neutralized with sodium thiosulfate if the water is chlorinated).
    dose for 20 to 24,000 chicks.
    Application to: D0 - D1 - D2 - D10 - D20 - D30 - D40
    Solution preparation procedure:
    Let the product thaw at room temperature between 15 and 25 ° C until complete liquefaction (approx. 1h ± 15 minutes) or 5 minutes in a 30 ° C water bath.
    Fill the tank with approximately 100L of water for applications up to D10 and 300L for others (if the water is treated with chlorine, add sodium thiosulfate up to 16 mg / L of water. other water treatment is performed, stop it on treatment days).
    Remove the cap and remove the stopper from the vials
    Pour the two bottles of the product into the water
    Mix to homogenize the product
    If an antibiotic treatment is carried out when the chicks arrive, delay the water circuit treatments to two days after the end of the antibiotic.
    d. Results
    Figures 4-10 show the results for lameness, mortality, treatment with antibiotics, burnt tarsus in males, and food consumption.
    Figures 4 and 5 show a significant decrease in the number of animals with burnt tarsi in males (Figure 4) or females (Figure 5) during the triple treatment with the composition according to the invention, compared to the control sample .
    FIGS. 6 and 7 show that the animals treated with the composition according to the invention according to the three modes of application significantly need less antibiotics. There is a trend of 12 ppm / kg bodyweight of antibiotic administered less in the test batches compared to the controls, ie a decrease of 39% in the consumption of antibiotics.
    The males have a greater weight after treatment with the composition according to the invention (FIG. 8), just like the females before the 1st slaughter (FIG. 9).
    Finally the mortality is reduced in the animals having been treated with the compositions according to the invention compared to the control animals (FIG. 10)
    In conclusion, there is a significant benefit for the health of the animals, and their well-being during the simultaneous use of the composition in the environment, application to the animal and ingestion by drinking water.
    EXAMPLE 5 - Application in the environment, on animals and by inhalation:
    pigs
    The purpose of this test is to show whether a combined application in animal environment and inhalation is effective. So check against the previous example, if other types of applications can be performed such as inhalation instead of ingestion.
    The test was carried out in 2 different farms according to the following protocols.
    Breeding 1:
    Stadium Event Control treatment (placebo) PS test treatment PS + ENG test treatment PS +ENG Pre-emptying, disinfection cleaning (ND) then emptysanitary (VS) Yes Yes Yes PS N b pigs / room 180 180 180 Prlv * OJ Yes Yes Yes Pulv * OJ Placebo 2 doses / 1000 m 2 2 doses / 1000 m 2 Pulv J15 Placebo 1 dose / 1000 m 2 1 dose / 1000 m 2 Pulv J30 Placebo 1 dose / 1000 m 2 1 dose / 1000 m 2 Prlv J42 Yes Yes Yes ENG Prlv OJ Yes Yes Yes Pulv OJ Placebo Placebo 1 dose / 1000 m 2 Pulv J15 Placebo Placebo 1 dose / 1000 m 2 Pulv J45 Placebo Placebo 1 dose / 1000 m 2 Prlv J50 Yes Yes Yes Pulv J75 Placebo Placebo 1 dose / 1000 m 2 Prlv J90 Yes Yes Yes
    * Prlv = bacteriological samples, Pulv = Nebulization, ENG: fattening,
    PS: post weaning
    Breeding 2:
    Stadium Event Treatment (placebo) witness Trial treatmentPS + Eng PS + ENG Pre-drain, ND Yes Yes
    then VS PS Nb pigs / room 350 350 Prlv * OJ Yes Yes Pulv * OJ Placebo 2 doses / 1000 m 2 Pulv J15 Placebo 1 dose / 1000 m 2 Pulv J30 Placebo 1 dose / 1000 m 2 Prlv J42 Yes Yes ENG Prlv OJ Yes Yes Pulv OJ Placebo 1 dose / 1000 m 2 Pulv J15 Placebo 1 dose / 1000 m 2 Pulv J45 Placebo 1 dose / 1000 m 2 Prlv J50 Yes Yes Pulv J75 Placebo 1 dose / 1000 m 2 Prlv J90 Yes Yes ABA Slaughter J100(Snouts) Yes Yes
    * Prlv = bacteriological samples, Pulv = Nebulization, ENG: fattening, PS: post weaning, ABA: Slaughter
    In this example, animals treated with the composition according to the invention according to the three modes of treatment are compared with animals which have not received the composition of the invention (placebo).
    The application was carried out with a nebulizer in the presence of animals so the flora were applied to the surfaces of the rooms (E), to the animals (A) and in inhalation (I).
    Pigs taken at random from farm 2 were analyzed after slaughter by measuring 10 the deformations on snout sections.
    Different bacteriological samples were taken before the first application of flora in the different rooms to assess the quality of the cleaning and disinfection step and at the exit of the animals from the rooms after the different applications of flora to assess the pathogenic pressure.
    Results:
    The results are compiled in the following table:
    Nose notation (snout cut in slaughterhouse) Breeding 2 Placebo n = 20 PC Floran = 19 PC Note 0 6 12 Note 1 10 7 Note 2 3 0 Note 3 1 0 Note 4 0 0 Average 0.95 0.37 TB nasal condition 30% 63%
    TB nasal condition: very good nasal condition
    Note 0: absence of deviation of the partition and atrophy of the scrolls and cornets
    Note 4: deviation of the partition and atrophy of the scrolls and cornets
    The placebo sample (without composition) indicates 30% of individuals with a very good nasal anatomical state (absence of deviation of the nasal septum, absence of atrophy of the volutes and nasal cones).
    The test sample (pigs reared in the presence of barrier flora from the start of post-weaning to the end of breeding) reveals that twice as many (63%) of individuals record 10 a very good nasal anatomical state vs placebo.
    It appears that the application according to the invention has a significant effect on the health and well-being of animals.
    The bacteriological results show that the more well the phase of cleaning and disinfection of the rooms, the more the barrier flora showed better results on the inhibition of the pathogenic flora:
    Note on the quality of the disinfection cleaning phase(1 = excellent to 4 = absence) Note 1.3 Note1.9 Note2.2 Note2.4 Frequency of best barrier flora results on pathogenic flora vs placebo, at the end of the period 5/6(83%) 4/6(67%) 3/6(50%) 0/6(0%)
    conclusions
    This test also shows the importance of the environmental application, the effectiveness of which is improved when the cleaning and disinfection of the surfaces upstream of the application is well carried out. The interest of the product is measured both on animals (sanitary), and vis-à-vis the farmer (working conditions).
    EXAMPLE 6 - Application in the environment and on animals in comparison with environment, animal and ingested application: Pigs
    The purpose of this essay is to confirm the value of combining the three modes of application over the combination of only two of them.
    The breeding followed is a breeding of 700 sows breeder fattener in breeding selection (5 bands of 120 sows). The test was carried out on 2 strips within which 3 batches of 40 sows were made while respecting a homogeneous distribution of the rows of litter. The application protocol includes the spraying of a barrier flora in the environment, on the udders and perineum of sows as well as a probiotic administration to sows via the soup machine.
    Spraying protocols applied:
    Loti Lot 2 Lot 3 Lot 4 Bandaged 1 +2 1 1 +2 2 Antibiotic on piglets Yes Yes Yes no J-3 Environment and sows Environment and sows Environment and sows Environment and sows J-2 J-1 Udders and perineum Udders and perineum Udders and perineum Udders and perineum D0: Betlow J1 Udders and perineum Udders and perineum Udders and perineum Udders and perineum J4 Udders and perineum Udders and perineum Protocol equivalence A + E A + E A + E A + E + I
    All of the sows received a dose of probiotics from D-3 to D1 compared to parturition.
    In the second strip, all of the sows received shampoo before arrival at the maternity unit, unlike the first strip.
    Only the piglets in lot 4 did not receive an antibiotic at the time of care. The antibiotic used for all the other batches being very effective against positive flora, the inventors consider in this test that the protocol of these batches is equivalent to an application only environment and on the animal because the flora ingested by piglets n did not survive the antibiotic.
    The inventors are therefore interested here in the differences observed between lots 1 to 3 (A + E) with lot 4 (A + E + I).
    Results:
    Figures 11 and 12 show that with the three applications combined, the risk of developing diarrhea is lower and the risk of mortality also compared to an application that is only environmental and on animals.
    FIG. 13 shows that more huts containing piglets whose mothers received a shampoo before the application of flora are healthy from a digestive point of view compared to piglets whose sows have not received shampoo. Indeed, in the first case (strip 2) we observe at the bottom 78 boxes out of 120 with normal faeces while in the second case (strip 1), only 50 boxes out of 120.
    This result therefore also shows the importance of the application on the animal, favored by the shampoo of the sows for a better development of the positive flora, to increase the effectiveness of the product.
    conclusions
    In this example, the inventors have shown that the A + E + l application is more efficient than the A + E application. In addition, they have shown that if we favor the implantation of flora on the animal (application A better performed), we improve the prevention of diarrhea.
    The invention is not limited to the embodiments presented and other embodiments will be apparent to those skilled in the art.
    权利要求:
    Claims (10)
    [1" id="c-fr-0001]
    claims
    1. Bacterial composition comprising at least one strain of bacteria of the genus Bacillus, in particular of Bacillus subtilis and at least one strain of lactic bacteria, in particular at least one strain of Lactococcus lactis, for its use in the context of the prevention or treatment of pathogenic infections affecting the health of terrestrial farm animals, said composition being in a form adapted to be:
    at. Ingested or inhaled by said land animals, and
    b. is
    b.1. spread in the farming environment of said terrestrial farm animals, b.2. applied to said land animals,
    b.3. applied in the culture medium of said terrestrial farm animals and applied to said terrestrial farm animals.
    [2" id="c-fr-0002]
    2. Composition for its use according to claim 1, in which the bacterial composition comprises:
    - at least one of the following three Bacillus subtilis strains: NOL01, NOL02, NOL03, said strains being respectively deposited at the CNCM under the number CNCM I - 4606, CNCM I-5043 and CNCM I - 4607, and
    - at least the strain of lactic bacteria: Lactococcus lactis spp lactis 1 strain NOL11, said strain being deposited at the CNCM under the number CNCM I -4609.
    [3" id="c-fr-0003]
    3. Composition for its use according to claim 1 or claim 2, said composition comprising from 10 4 to 10 11 bacterial colonies of Bacillus and from 10 4 to 10 11 bacterial colonies of lactic bacteria, the bacterial cottonies being in grams or in milliliters of composition.
    [4" id="c-fr-0004]
    4. Composition for its use according to any one of claims 1 to 3, wherein said at least one strain of Bacillus is in sporulated form and / or in vegetative form.
    [5" id="c-fr-0005]
    5. Composition for its use according to any one of claims 1 to 4, wherein said composition in a form suitable for being inhaled or ingested and said composition in a form suitable for being spread in the breeding environment of said breeding animals terrestrial and / or in a form suitable to be applied to said terrestrial farm animals, are used simultaneously, separately or spread over time.
    [6" id="c-fr-0006]
    6. Composition for its use according to any one of claims 1 to 5, wherein said composition is in a form suitable for being spread and / or applied prior to ingestion or inhalation by said land animals.
    [7" id="c-fr-0007]
    7. Composition for its use according to any one of claims 1 to 6, in which said composition consists essentially of the four bacterial strains from the group consisting of the three Badllus subtilis strains NOL01, NOL02, NOL03, said strains being respectively deposited at the CNCM under the number CNCM I - 4606, CNCM I-5043 and CNCM I - 4607 and the strain Lactococcus lactis spp lactis 1 strain NOL11, said strain being deposited at the CNCM under the number CNCM
    I-4609.
    [8" id="c-fr-0008]
    8. Composition for its use according to any one of claims 1 to 7, wherein said composition is spread by m 2 in an amount of 10 5 to 10 11 bacterial colonies.
    [9" id="c-fr-0009]
    9. Composition for its use according to any one of claims 1 to 8, in which the environment is cleaned and disinfected and the terrestrial animals are washed before spreading in the breeding medium of said terrestrial breeding animals and / or the application to said terrestrial farm animals.
    [10" id="c-fr-0010]
    10. Pathogen infection prevention kit affecting the health and well-being of land-based animals including
    - a composition as defined in any one of the preceding claims, and
    - at least one biosecurity agent chosen from disinfectants, detergents, UV radiation, or even water vapor and / or essential oils.
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    同族专利:
    公开号 | 公开日
    EP3675885A1|2020-07-08|
    FR3073139B1|2021-05-21|
    WO2019092134A1|2019-05-16|
    US20200345794A1|2020-11-05|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    WO2013178947A1|2012-05-30|2013-12-05|Nolivade|Bacterial composition for the treatment of colibacillosis on farms, in particular poultry farms, and also drinking water containing such a bacterial composition|
    WO2014151837A1|2013-03-15|2014-09-25|Bayer Cropscience Lp|A bacillus subtilis strain for use in maintaining or improving the paw health of poultry|
    FR2758051B1|1997-01-06|1999-12-17|Cobiotex|METHODS FOR SYSTEMATICALLY ERADICATING THE CARRYING OF PATHOGENS BY ANIMALS AND COMPOSITIONS USED THEREIN|
    RU2546880C2|2013-06-03|2015-04-10|Федеральное государственное бюджетное учреждение науки Институт проблем экологии и эволюции им. А.Н. Северцова Академии Наук|Method of probiotic feed preparation for farm animals|
    CN103773721A|2013-12-31|2014-05-07|陕西万源生物农业科技有限公司|Composite probiotics for raising pigs|
    CN104307012B|2014-11-05|2017-08-11|双胞胎（集团）股份有限公司|A kind of special deodorant of livestock and poultry farm and its application|FR3105730A1|2019-12-31|2021-07-02|Nolivade|Composition for the treatment of mammalian pathologies|
    FR3106724A1|2020-01-31|2021-08-06|Nolivade|Composition for improving insect production|
    FR3106725A1|2020-01-31|2021-08-06|Nolivade|Composition of microorganisms for breeding insects|
    FR3110344A1|2020-05-20|2021-11-26|Nolivade|Decontamination process|
    法律状态:
    2019-05-10| PLSC| Publication of the preliminary search report|Effective date: 20190510 |
    2019-11-29| PLFP| Fee payment|Year of fee payment: 3 |
    2020-11-27| PLFP| Fee payment|Year of fee payment: 4 |
    2021-11-29| PLFP| Fee payment|Year of fee payment: 5 |
    优先权:
    申请号 | 申请日 | 专利标题
    FR1760495A|FR3073139B1|2017-11-08|2017-11-08|COMPOSITION IMPROVING ANIMAL WELFARE|
    FR1760495|2017-11-08|FR1760495A| FR3073139B1|2017-11-08|2017-11-08|COMPOSITION IMPROVING ANIMAL WELFARE|
    US16/762,260| US20200345794A1|2017-11-08|2018-11-08|Composition for improving the well-being of animals|
    EP18807884.4A| EP3675885A1|2017-11-08|2018-11-08|Composition for improving the well-being of animals|
    PCT/EP2018/080666| WO2019092134A1|2017-11-08|2018-11-08|Composition for improving the well-being of animals|
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