• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    The composition, pharmaceutical or probiotic, comprises at least one strain of Lactobacillus intestinal tropism, selected from the species Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus casei, Lactobacillus paracasei and Lactobacillus rhamnosus, for use to promote juvenile growth in case of malnutrition . These include malnutrition characterized by protein deficiency. The strains can be selected on a Drosophila model and / or on a mouse model. The invention also relates to a method of probiotic treatment using this composition.
    公开号:FR3020949A1
    申请号:FR1454422
    申请日:2014-05-16
    公开日:2015-11-20
    发明作者:Francois Leulier；Gilles Storelli；Martin Schwarzer
    申请人:ECOLE NORM SUPERIEURE LYON；Centre National de la Recherche Scientifique CNRS；Universite Claude Bernard Lyon 1 UCBL；Ecole Normale Superieure de Lyon；
    IPC主号:
    专利说明:

    [0001] The present invention relates to a pharmaceutical or probiotic composition for promoting juvenile human and animal growth in case of malnutrition. This composition comprises as active principle or ingredient at least one bacterium intestinal tropism, preferably a lactic acid bacterium. Described as "an additional organ", the intestinal microbial community (or intestinal microbiota) plays a key beneficial role for the host by exerting many biological functions, such as aid in the efficiency of digestion, substrate metabolism , the fight against pathogens, or the establishment and homeostasis of immune responses. Imbalances between different intestinal bacterial populations have sometimes deleterious repercussions, leading to the development of various pathologies such as chronic inflammatory diseases or metabolic disorders including obesity or type 2 diabetes. These imbalances are also potentially involved in the development of cancers and the establishment of behavioral syndromes. Maintaining the balance of the gut microbiota is therefore essential: influencing its composition and / or its activity would be a major asset for the treatment of the aforementioned syndromes. In this spirit, the idea of using so-called "probiotic" bacterial strains to intervene on pathologies influenced by the microbiota finds its meaning. Defined in 2001 by the World Health Organization (WHO) and the Food and Agriculture Organization of the United Nations (FAO), probiotics are "living microorganisms that, when ingested in sufficient quantity, have positive effects on health, beyond the traditional nutritional effects. The present invention is in turn in the restoration of conditions for optimizing juvenile growth or restore normal juvenile growth in a context of malnutrition. The object of the present invention is therefore to propose novel compositions for promoting juvenile growth or for restoring normal juvenile growth in subjects, human or animal, having been or being subjected to malnutrition, in particular undernourishment and / or bad assimilation. Another object of the invention is to provide such compositions, based on the use of bacterial strains having an intestinal tropism, including commensal strains, or acceptable as a probiotic. Another object of the invention is to provide such compositions that can also be part of a therapeutic context.
    [0002] Yet another object of the invention is to provide a method of probiotic or therapeutic treatment. Yet another object of the invention is to provide such compositions and methods for supporting a health and / or nutritional claim in accordance with the legislation in force, including European legislation.
    [0003] The invention is based on the fact that certain strains of bacteria having an intestinal tropism in an animal species have an effect favoring or restoring the juvenile growth in a subject of the same species or of another species which is subjected to malnutrition, in particular, undernutrition or poor assimilation. It has thus been demonstrated that bacterial strains of the genus Lactobacillus were able to promote juvenile growth both in a Drosophila model and in a mouse model coupled with deficient nutritional diets, and furthermore a link could be established between these results. in mice and an increase in serum IGF-1 titre in mice treated under these conditions with these bacteria.
    [0004] The subject of the present invention is therefore a composition comprising at least one strain of bacterium having an intestinal tropism, in particular a lactic acid bacterium, for use in promoting juvenile growth in the event of malnutrition. The concept of malnutrition includes under-nutrition, over-nutrition and mis-assimilation. The invention is particularly aimed at undernourishment and / or poor assimilation. It is preferably aimed at undernourishment. The indication of the composition may be therapeutic or health, as a medicament, or nutritional, as a probiotic. By bacterium having a "intestinal tropism" is meant a bacterium with the ability to pass the gastric barrier and which is able to persist in the intestine.
    [0005] According to an advantageous characteristic of the invention, the bacterium can promote the production of IGF-1 in humans or animals which is treated with the composition according to the invention. The invention particularly provides strains of bacteria belonging to the following families: Lactobacillaceae, Streptoccaceae, Enterococcaceae, Leuconostocaceae, Bifidobacteriaceae. According to one embodiment, the invention uses one or more strains of the genus Lactobacillus, in particular one of the following species, Lactobacillus delbrueckii, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus casei, Lactobacillus paracasei, Lactobacillus rhamnosus. More particularly, they are bacteria belonging to the species Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus casei, Lactobacillus paracasei, Lactobacillus rhamnosus. In one embodiment, the bacterial strain is selected from L. plantarum WJL, L. casei ATCC 393, L. casei L919, L. paracasei ATCC25302, L. paracasei Shirota, L. fermentum ATCC9338, L. rhamnosus L900, L rhamnosus L908, L. rhamnosus GG. In a specific embodiment, they are bacteria of the species Lactobacillus plantarum, for example the strain WJL. According to a modality, the compositions according to the invention comprise at least one bacterial strain chosen from these groups, which has the required properties and makes it possible to promote or restore a juvenile growth despite the episode of malnutrition. Of course, the composition according to the invention may comprise more than one strain of bacteria meeting the needs of the invention. In particular, the composition comprises 2 or more of these bacterial strains, chosen from the same species or from different species.
    [0006] According to an interesting embodiment, the bacterium is a L. plantarum. A suitable strain is L. plantarum WJL (Eun-Kyoung Kim et al., Genome Announcements, November / December 2013, Vol.1, No. 6 e00937-13, GenBank AUTE00000000, Lactobacillus plantarum WJL, whole genome shotgun sequencing project). This strain WJL was initially isolated and can be isolated from Drosophila (JH Ryu et al., Science 2008, 319: 777-782). Other examples of suitable strains are: L. casei ATCC 393, L. casei L919 (Koryszewska-Baginska A. et al., September 26, 2013, Genome Announc), L. paracasei ATCC25302, L. paracasei Shirota (Yuki N et al., Int J Food Microbiol, April 1, 1999, 48 (1): 51-7), L. fermentum ATCC9338, L. rhamnosus L900 (Aleksandrzak-Piekarczyk T. et al., Genome Announc, August 15, 2013) , L. rhamnosus L908 (Koryszewska-Baginska A. et al., February 20, 2014, Genome Announc), L. rhamnosus GG (Kankainen M. et al., Proc Natl Acad Sci USA, October 6, 2009). The present invention thus brings to the art the teaching that bacterial strains with intestinal tropism promote juvenile growth, particularly in malnourished (human or animal) patients. But the invention is not limited to this teaching, it also gives the skilled person to safely determine the bacterial strains useful for the invention.
    [0007] Different criteria can be the basis of tests, being used alone or in combination. These criteria include serum IGF-1 level in the model animal (eg mice), larval growth in Drosophila melanogaster, model mouse growth as illustrated, for example, by femur size or animal size. On the basis of these criteria or similar criteria, it is possible for those skilled in the art to develop tests comparing individuals raised in the presence or absence of the bacteria to be tested under conditions of malnutrition. The invention provides a further by proposing axenic animal models and the breeding of animals in the presence or in the absence of the bacteria or bacteria to be tested making it possible to concentrate the test on the analysis of the intrinsic properties of the strain (s). tested and thus to overcome the effect that the resident gut microbiota of conventional animals could have in the context of the functional test. This results in increased predictivity of the test as to the functional potential of the strain tested.
    [0008] By "axenic" organism is meant an organism (e.g. Drosophila, mouse) raised in an environment free from microorganisms and therefore devoid of intestinal flora. The term "monoxenic" organism means an organism (e.g. Drosophila, mouse) associated axen high in the presence of a single microorganism and therefore carrying this unique microorganism as intestinal flora. According to one characteristic of the invention, the bacterial strains according to the invention are characterized by the fact that they respond positively to, or have been selected using, an assay in which the growth of larvae derived from axenic drosophilae is compared with a nutrient medium deficient yeast (and therefore protein) compared to a conventional nutrient medium. The conventional laboratory nutritional media generally comprise at least a plant meal, typically corn, inactivated yeast, typically brewer's yeast or baker's yeast (Saccharomyces cerevisiae), agar and water. An inactivated yeast deficient medium does not allow optimum postembryonic (juvenile) development of Drosophila larvae, as reported in G. Storelli et al., Eye Metabolism 14, 403-414, 2011. A group of The embryos are further inoculated with the bacterium to be tested, allowing the larvae emerging from the embryos to be associated with the bacteria to be tested. It can therefore be detected whether this bacterium is likely to promote or allow juvenile growth despite a nutrient medium deficient in yeast. The Drosophila test is typically conducted in the following manner: a batch of embryos derived from Drosophila melanogaster (e.g. Drosophila yw, or other so-called "wild-type" strain) relatives is available; at day 1: these embryos are divided into at least 2 groups (40 embryos, made into a triplicate for a total of 120 embryos per group) on a nutritional medium comprising maize meal of agar and water, deficient in yeast, one of the embryo groups is further inoculated with a suspension of approximately 108 CFU of the test bacteria in a saline buffer (eg PBS), which forms the monoxenic group, the other group forming the group axenic, the culture is conducted at about 25 ° C until J7; typically the culture medium consists, for 1 liter of medium, of 7.14 g of agar, 80 g of corn flour and 6 g of inactivated yeast, the medium is cooked in boiling water for 10 min, then cooling ; typically, the same medium, not deficient in inactivated yeast, contains 50 g of inactivated yeast; at day 7: at least 60 larvae of Drosophila from each group obtained from the inoculated or non-inoculated embryos (respectively monoxenic larvae and axenic larvae) are recovered, they are applied a rapid heat shock (eg 5 seconds, typically the larvae are placed for 5 seconds on a plate heated to 100 ° C, it is a question of killing the larvae without deforming them to allow the measurement of their size); the mean length of the larvae of each group is determined, and the averages obtained are compared; the bacterial strain tested is considered to respond positively to the test if the mean length of the larvae of the monoxenic group is greater than the mean of the length of the larvae of the axenic group with a value of p less than or equal to 0.001, preferably less than or 0.0001 in the Mann-Whitney Statistical Test, performed on the entire larval size dataset of both groups. The subject of the present invention is therefore a composition comprising at least one bacterial strain having an intestinal tropism, in particular a lactic acid bacterium, for use in promoting juvenile growth in the event of malnutrition, in which the strain of bacteria responds positively to the test on Drosophila as described above. By way of examples, mention may be made of the following strains: L. plantarum WJL, L. casei ATCC 393, L. casei L919, L. paracasei ATCC25302, L. paracasei Shirota, L. fermentum ATCC9338, L. rhamnosus L900, L rhamnosus L908, L. rhamnosus GG. Other strains can be identified among intestinal tropism bacteria and in particular among the species mentioned above. In one embodiment of the invention, strain WJL is used as a reference strain to identify and select bacterial strains having a "marked" effect on juvenile growth, namely an effect close to that of this strain WJL (effect not significantly different from the strain WJL), or a "strong" effect on juvenile growth (effect significantly greater than the strain WJL).
    [0009] For this purpose, the Drosophila test (including 60 larvae per condition) is applied to the strain WJL and to the strain to be tested (in parallel, preferably, or else reference data previously generated for the strain WJL can be obtained by example the data presented in the examples). The averages obtained for the two strains are then compared. The bacterial strain tested is considered to be a marked effect strain if the mean larval length of the monoxenic group is not significantly different from the average larval length of the WJL group with a p value greater than 0.001. in the Mann-Whitney statistical test, performed on the entire dataset. The effect is strong if said average for the strain to be tested is significantly greater than the average for the strain WJL, which is the case when the value of p of the statistical test is less than or equal to 0.0001. The effect is qualified as intermediate if said average for the strain to be tested is significantly lower than the average for the strain WJL, which is the case when the value of p of the statistical test is less than or equal to 0.0001.
    [0010] The subject of the present invention is therefore a composition comprising at least one bacterial strain having an intestinal tropism, in particular a lactic acid bacterium, for use in promoting juvenile growth in the event of malnutrition, in which the strain of bacteria has a marked effect or strong on growth in the Drosophila model as described above. By way of examples, mention may be made of the following strains: L. plantarum WJL, L. casei ATCC 393, L. casei L919, L. paracasei ATCC25302, L. paracasei Shirota, L. fermentum ATCC9338, L. rhamnosus L900, L rhamnosus L908, L. rhamnosus GG. Other strains can be identified among intestinal tropism bacteria and in particular among the species mentioned above.
    [0011] According to the invention, it is also possible to determine whether a strain of intestinal tropism bacterium can promote growth in case of malnutrition, using an axenic mouse model that allows monitoring of skeletal growth of mice in the presence of the disease. bacterium to be tested in comparison with the absence of microbiota and / or with the presence of a strain of reference bacterium. This model can be used as first-line therapy or on strains that have responded positively to the Drosophila test. According to one characteristic of the invention, the bacterial strains according to the invention are characterized by the fact that it responds positively to the following skeletal growth test: from the same mouse line (typically Balb / c mice) establishes an axenic parent mouse line and a monoxenic parent mouse line (associated with the bacterium to be tested), and juveniles are produced that are reared with the parents in a conventional nutritional medium until they are weaned (on D21); to form the group of monoxenic juveniles, we use parents mono-associated with the bacterial strain to be tested, on day 21: we have 8 weaned juveniles from each of these two lines, forming the monoxenic group and the axenic group, and they are raised in a nutritional diet deficient in protein (8%) and lipids (2.5%); typically, the conventional nutritional diet comprises 23% of proteins and 5% of lipids, at 56: the mean size of the mice is determined for a group considered by measuring the muzzle tip at the base of the tail of each individual; another possible measure consists in sacrificing the individuals, taking the femurs and measuring them, the lactic acid bacteria strain being considered as responding positively to the test if the average size of the individuals and / or femurs of the monoxenic group is greater than the average size, respectively, of individuals and / or femurs, of the axenic group, with a p-value of less than or equal to 0.05, in the Tukey statistical test. The subject of the present invention is therefore a composition comprising at least one bacterial strain having an intestinal tropism, in particular a lactic acid bacterium, for use in promoting juvenile growth in the event of malnutrition, in which the strain of bacteria responds positively to the test of skeletal growth on mice. The Drosophila test allows for a faster screening, so that usually the Drosophila test is also available, and the bacterial strain responds positively to Drosophila and skeletal growth tests. By way of example, mention may be made of the following strain: L. plantarum WJL. Other strains can be identified among the intestinal tropism bacteria and in particular among the species and strains mentioned above, in particular among the L. casei ATCC 393 strains, L. casei L919, L. paracasei ATCC25302, L. paracasei Shirota, L. fermentum ATCC9338, L. rhamnosus L900, L. rhamnosus L908, L. rhamnosus GG. In one embodiment of the invention, strain WJL is used as a reference strain to identify and select bacterial strains having a "marked" effect on juvenile growth, namely an effect close to that of this strain. WJL (effect not significantly different from strain WJL), or a "strong" effect on juvenile growth (effect significantly greater than strain WJL). To do this, the mouse test (including 8 mice per condition) is applied to the strain WJL and to the strain to be tested (in parallel, preferably, or else reference data previously generated for the strain WJL can be obtained by example the data presented in the examples). The averages obtained for the two strains are then compared. The bacterial strain tested is considered to be a marked-effect strain if the mean mean size of individuals and / or femurs of the monoxenic group is not significantly different from the corresponding average for the WJL group with a p-value. greater than 0.05 in the Tukey statistical test. The effect is strong if said average for the strain to be tested is significantly greater than the average for the strain WJL, which is the case when the value of p of the statistical test is less than or equal to 0.05. The effect is described as intermediate if said average for the strain to be tested (which has been qualified with respect to the axenic mice in the previous test) is significantly lower than the average for the strain WJL, which is the case when the value p of the statistical test is less than or equal to 0.05. The composition according to the invention will preferably comprise a strain of bacteria having such a marked or strong effect.
    [0012] The bacterial strains according to the invention can be characterized by the fact that they have a positive impact on the level of serum IGF-1. Thus it was possible, on the basis of an axenic mouse model, to show that mice reared in a medium with a low protein content but in the presence of the bacterium (monoxenic mice) had a higher growth rate and at the same time, a higher level of serum IGF-1, compared to these same mice raised with this protein-deficient medium and in the absence of the bacterium (axenic mice). This makes it possible to propose a test for determining whether a strain of bacteria has the potential to increase the serum level of IGF-1, which test can be applied in combination with a skeletal growth test in order to specify or refine the effect of the strain in this context of juvenile growth, or in first intention. In this case, the bacterial strains according to the invention are characterized by the fact that they respond positively to the following serum IGF-1 level test: from the same mouse line (typically Balb / c mice) an axenic parent mouse line and a monoxenic parent mouse line (associated with the bacterium to be tested) are established, and juveniles are produced that are raised with the parents in a conventional nutritional medium until they are weaned (at D21) ; to form the group of monoxenic juveniles, we use parents mono-associated with the bacterial strain to be tested, on day 21: we have 8 weaned juveniles from each of these two lines, forming the monoxenic group and the axenic group, and they are raised in a nutritional diet deficient in protein (8%) and lipids (2.5%); typically, the conventional nutritional diet comprises 23% protein and 5% lipid, at which time juveniles of each group are taken from blood and the average serum level of IGF-1 is determined for each group; this measurement of the serum level of IGF-1 is preferably carried out on diluted serum (1:25); commercial IGF-1 ELISA kits are preferably used according to the manufacturer's instructions, the lactic acid bacteria strain is considered to respond positively to the test if the average serum IGF-1 level of the monoxenic group is higher at the average serum level of the axenic group with a p-value less than or equal to 0.05 in the Tukey statistical test.
    [0013] The subject of the present invention is therefore a composition comprising at least one bacterial strain having an intestinal tropism, in particular a lactic acid bacterium, for use in promoting juvenile growth in the event of malnutrition, in which the strain of bacteria increases the level of Serum IGF-1. These include a strain of bacteria that responds positively to the test on mice as described above. The subject of the present invention is also a composition comprising at least one bacterial strain having an intestinal tropism, in particular a lactic acid bacterium, for use in promoting juvenile growth in the event of malnutrition, in which the strain of bacteria responds positively to the test of skeletal growth in mice and increases the serum level of IGF-1 in these same mice, particularly said strain also positively responds to the test on Drosophila. By way of example, mention may be made of the following strain: L. plantarum WJL. Other strains can be identified among the intestinal tropism bacteria and in particular among the species and strains mentioned above, in particular among the L. casei ATCC 393 strains, L. casei L919, L. paracasei ATCC25302, L. paracasei Shirota, L. fermentum ATCC9338, L. rhamnosus L900, L. rhamnosus L908, L. rhamnosus GG. In one embodiment of the invention, the WJL strain is used as a reference strain to identify and select bacterial strains having a "marked" effect on the serum IGF-1 level, namely an effect close to that of this strain WJL (effect not significantly different from the strain WJL), or a "strong" effect on the serum level of IGF-1 (effect significantly greater than the strain WJL). To do this, the mouse test (including 8 mice per condition) for measuring the serum level of IGF-1 is applied to the strain WJL and to the strain to be tested (in parallel, preferably, or else data may be available previously generated for the strain WJL, for example the data presented in the examples). The serum IGF-1 mean values obtained for the two strains are then compared. The bacterial strain tested is considered to be a marked effect strain if the average IGF-1 level of the monoxenic group is not significantly different from the mean for the WJL group with a p value greater than 0.05 , in Tukey's statistical test. The effect is strong if said average for the strain to be tested is significantly greater than the average for the strain WJL when the value of p is less than or equal to 0.05. The effect is described as intermediate if said average for the strain to be tested (previously qualified on the previous axenic test) is significantly lower than the average for the strain WJL when the value of p is less than or equal to 0.05. The composition according to the invention will therefore preferably comprise a bacterial strain having such a marked or strong effect, and in particular this bacterial strain has a marked or strong effect on both skeletal growth and serum IGF level. -1. The composition according to the invention, comprising at least one bacterial strain according to the invention, is suitable for use as a medicament or probiotic for promoting juvenile growth in case of malnutrition, especially with protein deficiency. According to a characteristic of the invention, the subject to be treated, malnourished or having been malnourished, may have a serum level of IGF-1 lower than that found in individuals of the same age and sex and not malnourished. The composition is usable in humans from birth to puberty. The composition can be used in livestock (cattle, sheep, goats, pigs, avian), pets (dog, cat) and sport (horse, camel, camel), between weaning and sexual maturity.
    [0014] The composition contains an amount of from 10 5 to 10 12, especially from 10 6 to 10 12, preferably from 10 8 to 10 9 bacterial colony forming cells (CFU) according to the invention, per gram of composition. The term CFU means "colony forming units" according to the English technical expression. Per gram of composition is preferably meant the pharmaceutical composition or the probiotic composition formed of the bacteria, co-ingredients, and excipients or vectors. By bacterial cells is meant a single strain of bacteria according to the invention or a mixture of at least two bacteria, according to the invention.
    [0015] The composition comprises the lactic acid bacteria or bacteria in living form. It can be a bacterial suspension, which can be frozen and thawed before use, or a lyophilized powder, which can be used as is or after recovery in a suitable vehicle. It can then comprise a conventional freeze-drying excipient. The composition may be a form of oral administration (for example powder, capsule, tablet) possibly in a gastro-protected form to pass the stomach and release the bacteria in the intestine. The invention also relates to a method of probiotic treatment or therapeutic treatment for promoting juvenile growth in case of malnutrition, including administration to a patient, human (from birth to puberty) or animal (from weaning to sexual maturity), who needs it, that is to say, malnourished or having undergone a period of malnutrition, in particular with protein deficiency, of a composition according to the invention. The patient in need may have serum IGF-1 levels lower than those seen in individuals of the same age and gender and not malnourished. As indicated, the method can be a therapeutic treatment or a probiotic treatment (or nutritional supplement). The method comprises administering a sufficient amount of a composition as described above. The amount and frequency of administration will depend in particular on the severity of the malnutrition, the age and the condition of the patient. The method will include administration in one or more times, which may be staggered over the growth period of the subject (until puberty or sexual maturity), doses of composition according to the invention. Doses can be fractionated to facilitate administration, especially depending on the age of the patient. The frequency of administration is in particular between a dose (single or fractional) every day and a dose every month. Typically, the frequency of administration will be between a dose (single or fractional) every day and a dose every week or every 2, 3, 4, 5 or 6 days. Each dose (single or fractional) represents in particular several grams to several tens of grams of composition. The method comprises in particular the administration of a composition comprising an amount of from 10 5 to 10 12, in particular from 10 6 to 10 12, preferably from 10 8 to 10 12 bacterial cells forming CFU colonies per gram of composition (by bacterial cells is meant a strain single bacterium according to the invention or a mixture of at least two bacteria, according to the invention). The method comprises in particular the administration of a composition comprising the lactic acid bacteria or bacteria in living form. This composition can be a bacterial suspension, which can be frozen and thawed before use, or a lyophilized powder, which can be used as it is or after recovery in a suitable vehicle. This composition may then comprise a conventional freeze-drying excipient, be a form of oral administration (for example powder, capsule, tablet) possibly in a gastro-protected form to pass the stomach and release the bacteria in the intestine, or still a rectal form (eg suppository). In one embodiment, the composition according to the invention, in particular the composition used in the treatment method, comprises at least one strain of bacteria which is not naturally present ("not-naturally occuring") in the species with which patient to be treated belongs. Thus, it may be a bacterium that is not naturally present in humans or animals. In this configuration, when there are several different bacteria, it is sufficient that one of them is not naturally present. On the other hand, this bacterium proves to have an intestinal tropism in the patient to be treated or in the corresponding species (human or animal according to the invention) and meets the definition of the active strains according to the invention. The invention also relates to a method for screening bacteria capable of promoting juvenile growth in case of malnutrition, using an axenic Drosophila model and / or an axenic mouse model. The method with the Drosophila model includes the following steps: a batch of embryos derived from Drosophila melanogaster (e.g. Drosophila strain yvv or other so-called "wild" strain) axenic; these embryos are distributed in at least 2 groups on a nutritional medium comprising a nutrient medium for yeast deficient drosophila, one of the groups of embryos is further inoculated with a suspension of the bacterium to be tested, which forms the group of monoxenic individuals; the other group forms the axenic group, the culture is carried out at the appropriate temperature (generally around 25 ° C), at the end of an appropriate breeding period (generally about 7 days), an equal number of the Drosophila larvae of each group from the embryos initially inoculated or not (axenic larvae and monoxenic larvae), the mean length of the larvae of each of the groups is determined, and the averages obtained are compared; the bacterial strain tested is considered to respond positively to the test if the mean length of the larvae of the monoxenic group is greater than the mean of the length of the larvae of the axenic group with a value of p less than or equal to 0.001, preferably less than or 0.0001 in the Mann-Whitney Statistical Test, performed on the entire larval size dataset of both groups.
    [0016] Preferably, the screening method incorporates the characteristics of the Drosophila test which has been described above. The method with the mouse model comprises the following steps: juveniles from two lines from the same mouse strain (typically Balb / c mice), namely an axenic parental mouse line and a monoxenic parental mouse line, are available. (associated with the bacterium to be tested), they are raised in nutritional diet deficient in proteins and preferably also in lipids, at the end of an appropriate breeding period, the average values of one or more parameters of each group, these parameters being related to growth (for example, weight gain, skeletal growth, for example by measuring the size of the individuals or the size of their femurs), and / or at the level of serum IGF-1, the strain of lactic acid bacterium is considered to respond positively to the test if the mean value of the measured parameter of the monoxenic group is greater than the mean value of the measured parameter of the focused group with a p-value less than or equal to 0.05 in the Tukey statistical test. Preferably, the screening method incorporates the characteristics of the mouse skeletal growth test or serum IGF-1 level described above.
    [0017] The screening method can also be a comparative test with a reference strain, for example the WJL strain, and this test then resumes the characteristics described above for Drosophila or mouse tests. The invention will now be described in more detail with the aid of embodiments of the invention taken as non-limiting examples and referring to the drawing in which: FIG. 1 is a graph showing the growth rate ( in cm / day) of mice from D28 to D56; GF: axenic mice; WJL: mice raised in the presence of L. plantarum strain WJL; NIZO 2877: mice bred in the presence of L. plantarum strain WJL NIZO 2877; * p <0.05; * p <0.001; Figure 2 is a graph showing femur size at J56 in GF, WJL or NIZO 2877 mice; * p <0.05; * "p <0.001; and FIG. 3 is a graph showing the serum levels (in ng / ml) of male mice at day 56, groups GF, WJL or NIZO 2877 * p <0.05; *** P <0.001. Example 1: Functional Screening of Bacteria on Monosene Drosophila: J-1: use of Axenic Adult Drosophila (D. melanogaster yw); they are placed in laying cages comprising a bottom (Petri dish type) on which is disposed a conventional nutritional medium; spawning is ensured by maintaining the adult population in the cage 1 night at 25 ° C. OJ: 6 pieces of the nutritional medium are cut out so as to collect 6 times 40 embryos, each piece is placed on a deficient nutritional medium contained in a Petri dish, three boxes are inoculated with sterile PBS 1x and the other three with a suspension of the bacterium in 1x PBS. Incubated for 7 days at 25 ° C. J7: We then recover from each box, the larvae that have developed there (the experiment is exploitable from 20 larvae per box, so we have hereinafter used groups of more than 60 individuals). The larvae undergo shock heat treatment (5 seconds on a plate heated to 100 ° C), then the size of the larvae is measured. This protocol was applied to different strains of bacteria, and the experiments and results are summarized in Table 1 below: Table 1. J7 larval sizes Statistical analysis Qualitative test interpretation (quantitative level (mm)) Mean (n> 60, mm) Standard deviation Comparison / Axenic animals Comparison / L. plantarum WJL Value of p (1) Statistical significance Value of p (1) Axial statistical significance 2,693 0,497 <0,0001 **** Lactobacillus plantarum WJL 3,644 0,883 <0,0001 **** SPM (labeled) Lactobacillus plantarum 3,398 0,750 <0.0001 **** 0.0001 *** SPM (intermediate) NIZ02877 Lactobacillus casei ATCC 393 4.391 0.802 <0.0001 **** 0.0721 ns SPM (labeled) Lactobacillus fermentum 4.133 0.824 <0.0001 * ***> 0.9999 ns SPM (labeled) ATCC 9338 Lactobacillus fermentum KLD 2,741 0.626> 0.9999 ns <0.0001 **** No effect on growth Lactobacillus fermentum LMG 3,287 0,799 0,1423 ns 0, 0015 ** No effect on growth Lactobacillus paracasei ATCC 3,987 0.524 <0.0001 **** 0.4258 ns SPM (labeled) 25302 Lactobacillus paracasei 8L23 2,682 0.482> 0.9999 ns <0.0001 **** No effect on growth Lactobacillus paracasei 3,407 0.779 <0.0001 **** 0.0984 ns SPM (labeled) Shirota Lactobacillus delbrueckii spp.bulgaricus ATCC 11842 2.540 0.599> 0.9999 ns <0.0001 **** No effect on growth Lactobacillus casei L919 4,150 0,712 <0,0001 **** 0,2966 ns SPM (labeled) Lactobacillus rhamnosus L900 3,920 0,572 <0,0001 ****> 0,9999 ns SPM (labeled) Lactobacillus rhamnosus L908 4,112 0,576 <0.0001 **** 0.1277 ns SPM (labeled) Lactobacillus rhamnosus GG 4.036 0.488 <0.0001 **** 0.5272 ns SPM (labeled) * Significantly higher * Significantly lower than SPM = Growth promoter strain to the reference value the reference value (1) Statistical test used: Mann-Whitney test. Indications on the validation criteria of the phenotypic test "larval growth": The strain must fulfill at least criterion 1 to be validated and the extent of its activity can be qualified according to criterion 1 + 2.
    [0018] Criterion 1 qualitative = growth promoter strain with respect to the axenic condition: A statistical test is applied to all the length values of the associated individuals in relation to the values of the axenic individuals (60 larvae at least per condition). A p-value of less than 0.001 (Mann-Whitney test) qualifies the strain as a "growth promoter". In this case the following strains are called growth promoter: Lactobacillus plantarum WJL Lactobacillus plantarum NIZ02877 Lactobacillus casei ATCC 393 Lactobacillus fermentum ATCC 9338 Lactobacillus paracasei ATCC 25302 Lactobacillus paracasei Shirota Lactobacillus casei L919 Lactobacillus rhamnosus L900 Lactobacillus rhamnosus l908 Lactobacillus rhamnosus GG The following strains have no growth promoting effect: Lactobacillus fermentum KLD Lactobacillus fermentum LMG Lactobacillus paracasei BL23 Lactobacillus delbrueckii spp.bulgaricus ATCC 11842 Criterion 2 Quantitative = extent of growth promotion effect (with criterion 1 validated beforehand): A test statistic is applied to all the length values of the individuals associated with the strain (s) tested compared to the values of individuals mono-associated with the L. plantarum strain WJL (60 larvae at least per condition). Three cases are presented: The p value of the statistical test is greater than 0.001 (Mann-Whitney test), the strain will be described as a "marked" effect on larval growth (effect similar to L. plantarum strain WJL) . In the present example the following strains have a marked effect: Lactobacillus casei ATCC 393 Lactobacillus fermentum ATCC 9338 Lactobacillus paracasei ATCC 25302 Lactobacillus paracasei Shirota Lactobacillus casei L919 Lactobacillus rhamnosus L900 Lactobacillus rhamnosus L908 Lactobacillus rhamnosus GG Lactobacillus plantarum WJL The p value of the statistical test is less than 0.001 (Mann-Whitney test), the strain will be described as an "intermediate" effect on larval growth if the average size of individuals is lower than those associated with L. plantarum WJL (case of strain L. plantarum NIZO 2877 in the example presented). The strain will be described as a "strong" effect if the average size of the individuals is greater than those associated with L. plantarum WJL (case not identified in the present example).
    [0019] Analysis of the "larval growth" phenotypic test example: - 14 strains of Lactobacillus were tested (Tables 1) to determine their potential for growth promotion using the "larval growth" test in Drosophila melanogaster. - These strains are available at the ATCC, at the Institut Pasteur in Paris, at the Institut Pasteur in Lille or published in the scientific literature and available from referent researchers in the publications mentioned (Table 2).
    [0020] Table 2: Public collection and / or publication with genome sequence Lactobacillus plantarum WJL Kim et al., Genome Announc. 21 November 2013, 1 (6) .Pil: e00937-13 Lactobacillus plantarum NIZO2877 NIZO (2877) Lactobacillus casei ATCC 393 ATCC (393) Lactobacillus fermentum ATCC 9338 ATCC (9338) Lactobacillus fermentum KLD Pasteur Institute of Lille (A5.20) Lactobacillus fermentum LMG Pasteur Institute of Lille (A5.16) Lactobacillus paracasei ATCC 25302 ATCC (25302) Lactobacillus paracasei BL23 Pasteur Institute of Lille (A3.6) and Mazé et al., J. Bacteriol., May 2010; 192 (10): 2647-8 Lactobacillus paracasei Shirota Pasteur Institute of Lille (A3.5) Lactobacillus delbrueckii spp.bulgaricus ATCC (11842) and van de Guchte M, et al., Proc.Natl Acad Sci USA June 13, 2006 Lactobacillus casei L919 Koryszewska-Baginska A et al., Genome Announc, September 26, 2013 Lactobacillus rhamnosus L900 Aleksandrzak-Piekarczyk T et al. Genome Announc, August 15, 2013 Lactobacillus rhamnosus Koryszewska-Baginska L908 A et al. Genome Announc, February 20, 2014 Lactobacillus rhamnosus GG ATCC (53103) and Kankainen M et al., Proc Natl Acad Sci USA, October 6, 2009 - The size of a minimum of 60 individuals per condition is studied at day 7 after association with the bacterial strain to be tested, then rearing juvenile individuals in nutrient deficient yeast. Table 1 shows the mean and standard deviation of these datasets. - The statistical analysis of these results makes it possible to qualify according to criterion 1 of the "larval growth test" 10 strains of different species of Lactobacilli. This selection highlights a strictly strain specific functional effect and not necessarily related to a given species of Lactobacillus. - According to criterion 2, the statistical analysis of the results makes it possible to qualify as "intermediate" the effect of the L. plantarum strain NIZO 2877, whereas the other 9 growth promoting strains have "marked" effects. Example 2 Functional Screening of Bacteria in Mice - Phenotypic Analysis: Study in Mice of the Juvenile Growth Promoting Effect of Two "Intermediate" and "Marked" Strains (L. plantarum NIZ02877 and L. plantarum WJL, respectively) identified by the "larval growth" test of Example 1. The male offspring (minimum 15 individuals) of three groups of individuals from the same axenic mouse colony was studied, the first group consists of axenic juveniles (GF group), the second in juveniles from parents mono-associated with the L. plantarum strain WJL (WJL group), and the third in juveniles from parents mono-associated with the L. plantarum strain NIZ02877 ( NIZO group 2877). The parents and juveniles are reared in a conventional nutritional medium until weaning of the juveniles (J21), then the weaned juveniles are raised in nutritional medium deficient in proteins and lipids (8% and 2.5% respectively) until J56. Conventional Nutritional Environment: 23% Protein and 5% Lipid Deficient Nutritional Environment: 8% Protein and 2.5% Lipid Three parameters illustrating the juvenile growth of these individuals were studied: (1) Size gain over a 28-day period following weaning (from D28 to D56), (2) the size of the femurs of a batch of individuals (at least 9 individuals) representative of the population tested at D56 and finally (3) the serum level at D56 of the IGF-1 growth in at least 8 individuals. Statistical analyzes were performed using the Tukey test using GraphPad software (GraphPad Prism 5.04, San Diego, USA); values of p <0.05 are considered significant. (1) Size gain: Mice were anesthetized by brief exposure to isoflurane to allow measurement of mouse size (snout at base of tail) at days 28 and 56.
    [0021] Table 3: growth rate (size taken per day in cm from D28 to D56) - Nb = number of GF WJL NIZO 2877 mice Mean Standard deviation Nb Mean Standard deviation Nb Mean Standard deviation Nb 0.007321 0.009878 0.031114 0.009376 17 0.016429 0.011071 25 These data are shown in Figure 1. (2) Femur size: The mice are sacrificed to J56, a femur is removed, released from the muscle and its length is measured at Vernier caliper. Table 4: Length of femurs in mm at day 56 in males GF WJL NIZO 2877 10.2 11.2 10.9 10.2 11.6 11 10.5 11.9 10.5 9.9 11.6 11, 2 10.4 12 11 11.4 11 10.2 12 11 10.2 11.7 11.2 9.8 12 11 11.9 11.6 11 These data are presented in Figure 2. (3) Titles in IGF-1 in serum: IGF-1 titers are measured on serum obtained from the blood of mice sacrificed at J56. Measurement is performed on diluted serum (1:25) using the Ready-Set-Go ELISA kit (eBioscience, USA), following the manufacturer's instructions. Table 5: Serum titre in 1G F-1 in pg / ml (double measurement from the ELISA plates) GF WJL NIZO 2877 4526.75 4343.75 8823.25 8606.5 7291.25 6998.75 2825.5 2614, 75 9258.5 9122.25 5128 5102 2667.5 2773 8471.25 8660.5 4343.75 4396 3455.25 3219.5 7025.25 7078.25 5232.75 5180.5 7131.5 6839.5 11063 9557, 3 8471.25 8282.5 2164.2 2057.55 15125 14409 4866.5 4761.75 3140.75 3035.75 10023 9372 6363.25 6152.25 2509 2456.15 11535 12026 7211.25 6680.5 2588.25 2614.75 4213.25 4161 4432.5 4601.3 4770.5 4517 4013.3 3930 4940.3 3846.5 3597.3 3846.5 These data are presented in Figure 3. The results illustrate a "marked" effect L. plantarum WJL strain on growth rate (p <0.001 compared to axenic condition) and an "intermediate" effect of L. plantarum strain NIZ02877 (p <0.05 value compared with axenic condition and value p <0.001 compared to L. plantarum WJL). These effects are confirmed with the "IGF-1 level" parameter (same ranges of p values of the different tests), then reinforced with the parameter "femur size" (p value <0.001 between axenic conditions and L. plantarum NIZO 2877) All of these results demonstrate by scientific evidence the juvenile growth promoter effect of certain strains of Lactobacillus also responding positively to the "larval growth" test. All public documents cited here are incorporated by reference. Similarly, those skilled in the art will be able to refer to these various documents and commercial strain deposits referred to herein. 15
    权利要求:
    Claims (14)
    [0001]
    REVENDICATIONS1. Composition comprising at least one strain of Lactobacillus intestinal tropism, selected from the species Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus casei, Lactobacillus paracasei and Lactobacillus rhamnosus, for use to promote juvenile growth in case of malnutrition.
    [0002]
    2. The composition according to claim 1, wherein the Lactobacillus strain is selected from L. plantarum WJL, L. case! ATCC 393, L. casei L919, L. paracasei ATCC25302, L. paracasei Shirota, L. fermentum ATCC9338, L. rhamnosus L900, L. rhamnosus L908, L. rhamnosus GG.
    [0003]
    3. Composition according to claim 1 or 2, wherein the strain of Lactobacillus responds positively to the following test: there is a batch of embryos of parents Drosophila melanogaster axenic; at Ji: these embryos are distributed in at least 2 groups (40 embryos, made in triplicate or 120 embryos per group) on a nutritional medium comprising maize flour, agar and water, deficient in yeast, one of the embryo groups is further inoculated with a suspension of approximately 108 CFU of the bacteria to be tested in a saline buffer, which forms the monoxenic group, the other group being the axenic group, the rearing of both groups at about 25 ° C until D7; at day 7: at least 60 larvae of Drosophila from each group are recovered, a thermal shock is applied to them; the mean length of the larvae of each group is determined, and the averages obtained are compared; the bacterial strain tested is considered to respond positively to the test if the mean length of the larvae of the monoxenic group is greater than the mean of the length of the larvae of the axenic group with a value of p less than or equal to 0.001, preferably less than or 0.0001 in the Mann-Whitney Statistical Test, performed on the entire larval size dataset of both groups.
    [0004]
    4. Composition according to claim 3, comprising a bacterial strain which, compared to the strain L. plantarum WJL, in the same test on Drosophila, has a strong effect with an average length of the larvae of the monoxenic group with this strain. significantly higher than average for the monoxenic group with the strain WJL, when the p-value is less than or equal to 0.0001 in the Mann-Whitney statistical test, performed on the set of data set of larval sizes of both groups.
    [0005]
    A composition according to any one of claims 1 to 4, wherein the Lactobacillus strain responds positively to the following test: from the same mouse line an axenic parental mouse line and a monoxenic parent mouse line are established. (associated with the bacterium to be tested), and juveniles are produced which are raised with the parents in a conventional nutritional medium until their weaning (D21) at D21: we have 8 weaned juveniles from each of these two lines , forming the monoxenic group and the axenic group, and raised in nutritional diet deficient in proteins (8%) and lipids (2.5%); at day 56: the average size of the mice for each group considered is determined by measuring the muzzle end at the base of the tail of each individual, or the femurs of the individuals of each group are taken and the we measure these; the lactic acid bacterium strain being considered to respond positively to the test if the average size of the individuals or femurs of the monoxenic group is greater than the average size of the individuals or femurs, of the axenic group, with a value of p less than or equal to 0 , 05 in Tukey's statistical test.
    [0006]
    6. Composition according to claim 5, comprising a strain of bacteria which, compared to the strain L. plantarum WJL, in this same mouse test, has a strong effect with an average size of the mice or their monoxenic group. with this bacterium strain significantly above the average for the monoxenic group with the WJL strain, when the p value is less than or equal to 0.05 in the Tukey statistical test.
    [0007]
    The composition according to any of claims 1 to 6, wherein the Lactobacillus strain responds positively to the following test: from the same line of axenic mice an axenic parental mouse line and a parental mouse line are established. monoxenics (associated with the bacterium to be tested), and juveniles are produced which are raised with the parents in conventional nutritional medium until their weaning (D21), D21: we have 8 weaned juveniles from each of these two lines, forming the monoxenic group and the axenic group, and raised in nutritional diet lacking protein (8%) and lipids (2.5%); at day 56: juveniles of each group are taken from the blood and the average serum level of IGF-1 for each group is determined; the lactic acid bacterium strain being considered as responding positively to the test if the average serum level of IGF-1 of the monoxenic group is greater than the average serum concentration of the axenic group with a p value of less than or equal to 0.05 in the statistical test of Tukey.
    [0008]
    8. A composition according to claim 7, comprising a strain of bacteria which, compared with the strain L. plantarum WJL, in this same mouse test, has a strong effect with a mean serum level of IGF-1 of the monoxenic group. with this bacterium strain significantly above average for the monoxenic group with strain WJL, when the p value is less than or equal to 0.05 in the Tukey statistical test.
    [0009]
    9. Composition according to any one of claims 1 to 8, for use as a medicament or probiotic for promoting juvenile growth in case of malnutrition characterized by a protein deficiency.
    [0010]
    10. Composition according to any one of the preceding claims, for use as a medicament or probiotic for promoting juvenile growth in case of malnutrition characterized by a serum level of IGF-1 below normal.
    [0011]
    11. A composition according to any one of the preceding claims, wherein the bacterium is a Lactobacillus plantarum.
    [0012]
    12. Composition according to any one of the preceding claims, containing from 105 to 1012 CFU of Lactobacillus responding positively to the test, per gram of composition.
    [0013]
    13. A method of screening bacteria capable of promoting juvenile growth in case of malnutrition, wherein the bacterial strain is subjected to the following test: there is a batch of embryos of parents of Drosophila melanogaster axenic; these embryos are placed in at least 2 groups on a nutritional medium comprising a nutrient medium for yeast deficient drosophila, one of the groups of embryos is further inoculated with a suspension of the bacteria to be tested, which forms the group of monoxenic individuals; the other group forms the axenic group, the culture is carried out at the appropriate temperature, at the time of a suitable breeding period, Drosophila larvae of each group are preferably recovered in equal numbers (axenic larvae and larvae). monoxenics), the mean length of the larvae of each group is determined, and the averages obtained are compared; the strain of bacterium tested is considered to respond positively to the tests; the mean length of the larvae of the monoxenic group is greater than the average of the length of the larvae of the axenic group with a p-value of less than or equal to 0.001, preferably less than or equal to 0.0001 in the Mann-Whitney test, performed on the entire dataset.
    [0014]
    14. A method for screening bacteria capable of promoting juvenile growth in the event of malnutrition, in which the bacterial strain is subjected to the following test: juveniles from two lines from the same strain of mice, namely line of axenic mice and a line of monoxenic mice, they are raised in nutritional diet deficient in proteins and preferably also in lipids; at the end of an appropriate breeding period, the average of the values of one or more parameters of each group is determined, these parameters being related to the level of serum IGF-1 or to growth, the strain of lactic acid bacterium is considered to respond positively to the test if the average level of IGF-1 in the monoxenic group is greater than the average rate of the axenic group with a p value of less than or equal to 0.05 in the Tukey test.20
    类似技术:
    公开号 | 公开日 | 专利标题
    Zagato et al.2014|Lactobacillus paracasei CBA L74 metabolic products and fermented milk for infant formula have anti-inflammatory activity on dendritic cells in vitro and protective effects against colitis and an enteric pathogen in vivo
    Zhang et al.2015|Impacts of gut bacteria on human health and diseases
    Endo et al.2013|Honeybees and beehives are rich sources for fructophilic lactic acid bacteria
    Stanley et al.2012|Intestinal microbiota associated with differential feed conversion efficiency in chickens
    JP2018048112A|2018-03-29|Bifidobacterium longum and functional gi disorders
    Sun et al.2013|Application of autochthonous Bacillus bioencapsulated in copepod to grouper Epinephelus coioides larvae
    EP3164144B1|2021-07-28|Lactobacillus composition allowing the stimulation of human and animal juvenile growth in cases of malnutrition
    Sumon et al.2018|Growth performance, digestive enzyme activity and immune response of Macrobrachium rosenbergii fed with probiotic Clostridium butyricum incorporated diets
    de La Banda et al.2010|Influence of two closely related probiotics on juvenile Senegalese sole | performance and protection against Photobacterium damselae subsp. piscicida
    EFSA Panel on Biological Hazards | et al.2018|Update of the list of QPS‐recommended biological agents intentionally added to food or feed as notified to EFSA 8: suitability of taxonomic units notified to EFSA until March 2018
    Thurlow et al.2019|Bacillus velezensis AP193 exerts probiotic effects in channel catfish | and reduces aquaculture pond eutrophication
    Deobagkar et al.2012|A study on bacteria associated with the intestinal tract of farmed yellow seahorse, Hippocampus kuda |: characterization and extracellular enzymes
    Bjornsdottir et al.2010|Selection of bacteria and the effects of bacterial treatment of Atlantic halibut | eggs and larvae
    de Assis et al.2016|Intestinal anti-inflammatory activity of goat milk and goat yoghurt in the acetic acid model of rat colitis
    Ren et al.2013|Lactobacillus planarum subsp. plantarum JCM 1149 vs. Aeromonas hydrophila NJ-1 in the anterior intestine and posterior intestine of hybrid tilapia Oreochromis niloticus♀× Oreochromis aureus♂: An ex vivo study
    CA2878130A1|2014-01-16|Novel strain of lactobacillus crispatus
    JP4565057B2|2010-10-20|Novel lactic acid bacteria with high ability to induce immunoglobulin A
    Landete et al.2018|Virulence and antibiotic resistance of enterococci isolated from healthy breastfed infants
    Shahi et al.2014|Recovery of Pseudomonas koreensis from eye lesions in golden mahseer, Tor putitora | in Uttarakhand, India
    Amoah et al.2020|Administration of probiotic Bacillus licheniformis induces growth, immune and antioxidant enzyme activities, gut microbiota assembly and resistance to Vibrio parahaemolyticus in Litopenaeus vannamei
    Khan2013|Probiotic microorganisms-identification, metabolic and physiological impact on poultry
    CA2893990C|2021-05-11|Probiotic strains for the treatment and/or prevention of diarrhea
    WO2018104336A1|2018-06-14|Lactic acid bacteria of use for treating metabolic diseases, especially type 2 diabetes
    FR3047640A1|2017-08-18|PROBIOTIC COMPOSITION FOR PROMOTING JUVENILE GROWTH IN ANIMAL BREEDING ANIMALS
    Adeshina et al.2019|Gut Microbiota and Innate Immune Response of Macrobrachium vollenhovenii Infected with Pseudomonas aeruginosa and Aeromonas hydrophila Fed Diets Supplemented with Lactobacillus acidophilus
    同族专利:
    公开号 | 公开日
    EP3164144B1|2021-07-28|
    DK3164144T3|2021-09-27|
    FR3020949B1|2018-02-09|
    CN106795482B|2021-03-12|
    US20170080034A1|2017-03-23|
    EP3164144A1|2017-05-10|
    PT3164144T|2021-09-24|
    ES2893866T3|2022-02-10|
    PL3164144T3|2022-01-17|
    WO2015173386A1|2015-11-19|
    CN106795482A|2017-05-31|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    WO2013057063A1|2011-10-18|2013-04-25|Nestec S.A.|Composition for use in the promotion of healthy bone growth and/or in the prevention and/or treatment of bone disease|
    WO2013191845A1|2012-06-20|2013-12-27|Mjn U.S. Holdings Llc|Dietary management of celiac disease and food allergy|
    US2735351A|1956-02-21|abrahamsen|
    AU2010247469B2|2009-05-11|2014-12-18|Société des Produits Nestlé S.A.|Lactobacillus johnsonii La1 NCC533 and immune disorders|
    WO2013057049A1|2011-10-18|2013-04-25|Nestec S.A.|Composition for use in brain growth and/or cognitive and/or psychomotor development|FR3047640A1|2016-02-12|2017-08-18|Ecole Normale Superieure Lyon|PROBIOTIC COMPOSITION FOR PROMOTING JUVENILE GROWTH IN ANIMAL BREEDING ANIMALS|
    FR3059678A1|2016-12-05|2018-06-08|Ecole Normale Superieure Lyon|LACTIC ACID BACTERIA USEFUL IN TREATING METABOLIC DISEASES, IN PARTICULAR TYPE 2 DIABETES|
    FR3070167B1|2017-08-17|2021-12-24|Ecole Normale Superieure Lyon|GROWTH-Stimulating LACTOBACILLUS STRAINS|
    CN113747906A|2019-08-26|2021-12-03|真共生株式会社|Biointroduction aid and method for using same|
    法律状态:
    2015-03-26| PLFP| Fee payment|Year of fee payment: 2 |
    2015-11-20| PLSC| Publication of the preliminary search report|Effective date: 20151120 |
    2016-02-25| PLFP| Fee payment|Year of fee payment: 3 |
    2017-03-16| PLFP| Fee payment|Year of fee payment: 4 |
    2018-05-22| PLFP| Fee payment|Year of fee payment: 5 |
    2019-05-22| PLFP| Fee payment|Year of fee payment: 6 |
    2020-05-22| PLFP| Fee payment|Year of fee payment: 7 |
    2021-05-20| PLFP| Fee payment|Year of fee payment: 8 |
    优先权:
    申请号 | 申请日 | 专利标题
    FR1454422A|FR3020949B1|2014-05-16|2014-05-16|COMPOSITION FOR PROMOTING HUMAN AND ANIMAL JUVENILE GROWTH IN CASE OF MALNUTRITION|
    FR1454422|2014-05-16|FR1454422A| FR3020949B1|2014-05-16|2014-05-16|COMPOSITION FOR PROMOTING HUMAN AND ANIMAL JUVENILE GROWTH IN CASE OF MALNUTRITION|
    PL15722225T| PL3164144T3|2014-05-16|2015-05-15|Lactobacillus composition allowing the stimulation of human and animal juvenile growth in cases of malnutrition|
    PT157222258T| PT3164144T|2014-05-16|2015-05-15|Lactobacillus composition allowing the stimulation of human and animal juvenile growth in cases of malnutrition|
    CN201580035168.9A| CN106795482B|2014-05-16|2015-05-15|Lactobacillus composition allowing to promote the juvenile growth of humans and animals in case of malnutrition|
    DK15722225.8T| DK3164144T3|2014-05-16|2015-05-15|Lactobacillus composition for stimulating juvenile growth in humans and animals by malnutrition|
    PCT/EP2015/060753| WO2015173386A1|2014-05-16|2015-05-15|Lactobacillus composition allowing the stimulation of human and animal juvenile growth in cases of malnutrition|
    ES15722225T| ES2893866T3|2014-05-16|2015-05-15|Composition of lactobacillus that allows to promote human and animal juvenile growth in case of malnutrition|
    US15/311,457| US20170080034A1|2014-05-16|2015-05-15|Lactobacillus composition allowing the stimulation of human and animal juvenile growth in cases of malnutrition|
    EP15722225.8A| EP3164144B1|2014-05-16|2015-05-15|Lactobacillus composition allowing the stimulation of human and animal juvenile growth in cases of malnutrition|
    [返回顶部]