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    • 专利摘要:
    Christensenella minuta strain and use of it. The present invention relates to the Christensenella minuta strain DSM 32891, to its cellular components, metabolites, and secreted molecules, and to compositions comprising the above products, as well as to the use of said strain for the prevention and/or treatment or alterations. mood or affective, such as depression. (Machine-translation by Google Translate, not legally binding)
    公开号:ES2763350A1
    申请号:ES201831153
    申请日:2018-11-28
    公开日:2020-05-28
    发明作者:Herranz Yolanda Sanz；DEL PULGAR VILLANUEVA EVA Mª GÓMEZ；Feliú Ana Agustí；LAGUNA Mª CARMEN CENIT
    申请人:Consejo Superior de Investigaciones Cientificas CSIC；
    IPC主号:
    专利说明:

    [0002] Christensenella minuta strain and use of it
    [0004] The present invention relates to the Christensenella minuta strain DSM 32891 and to its use for the prevention or treatment of mood or affective disorders, such as depression. The present invention falls within the field of therapeutic activity of pharmaceutical compositions or preparations, as well as within the field of food.
    [0006] BACKGROUND OF THE INVENTION
    [0008] Mood disorders, and especially depression, are one of the leading causes of disability worldwide. The total cost of mental disorders is estimated to be 798 billion, of which mood disorders represent a direct and indirect annual cost of about 118 billion [1]. These include major depression, typical or melancholic and atypical depression, pre- and postpartum depression, bipolar disorder, psychotic depression, dysthymia, depressive personality disorder, seasonal affective disorder, mood-induced mood disorder. substance abuse or drug use etc. Furthermore, the effectiveness of current therapies is quite limited. Approximately only 50% of antidepressant treatments are estimated to be effective; many patients are left with sub-clinical symptoms and others do not show any improvement.
    [0010] Depression is a complex pathology characterized by the presence of heterogeneous symptoms, suggesting the existence of different forms of depression or phenotypes (for example, typical depression characterized by stronger hyperactivity of the hypothalamic-pituitary-adrenal axis [HPA] and depression atypical characterized by increased metabolic dysregulation and increased appetite / weight) [2, 3]. However, little is known about the molecular mechanisms underlying these pathologies. Depression also shows high comorbidity with mental (for example, anxiety) and physical (for example, cardio-metabolic disorders, such as metabolic syndrome, diabetes and CVD), which worsen the course of the disease and reduce the response therapeutic and increase your risk of it.
    [0011] Epidemiological research in humans has revealed associations between alterations in the configuration of the intestinal microbiota (dysbiosis) and psychiatric disorders, such as mood or affective disorders and, among these, especially depression [4-9]. These associations between dysbiosis and depression are thought to be largely determined by environmental psychosocial factors (childhood trauma, work stress, lack of sleep) and lifestyle (poor diets, sedentary lifestyle, medication) as well as other characteristics of the individual. such as its genome, age, sex, and the presence of comorbidities [2, 10]. In animal models, increased stress-induced HPA axis response, a well-established risk factor for depression, causes intestinal dysbiosis; in turn, the dysbiotic microbiota contributes to behavioral and mood disturbances [11]. Animal studies also show that the specific configuration of the intestinal microbiota influences the stress response by aggravating or improving its neurochemical and behavioral consequences, through mechanisms that coordinate the dialogue of the immune, endocrine and nervous systems [8, 12, 13].
    [0013] This evidence suggests that the use of strategies aimed at modulating the composition and functions of the microbiota could be an alternative from the point of view of prevention and treatment of mood or affective disorders.
    [0015] The use of traditional probiotics (lactobacilli and bifidobacteria) for the treatment of depression has been shown to have different effects depending on the strain used. Bifidobacteria have possibly been the most widely used to assess their effectiveness against anxiety and depression; however, the results obtained have not always been conclusive and of sufficient magnitude [14,15]
    [0017] Beyond bifidobacteria, other bacterial species may be of interest for these applications, but observational studies that establish associations between different bacterial groups and depression, because they increase or decrease in subjects with it, are inconclusive. Thus, for example, Yu et al. (2017) [16], observed low proportions of the bacterial groups Marvinbryantia, Corynebacterium, Psychrobacter, Christensenella, Lactobacillus, Peptostreptococcaceae incertae sedis, Anaerovorax, Clostridialesincertae sedis and Coprococcus in mouse models with induced depression. However, Mironova et al. (2017) [17] published that the microbiota of patients with Parkinson's disease and moderate depression had a higher abundance of Christensenella minuta, Clostridium disporicum and Oscillibacter valericigenes compared to patients with Parkinson's disease and mild depression or absence of depression.
    [0019] For this reason, it is necessary to continue searching for more effective preventive and therapeutic strategies, based on new bacterial strains that are unambiguously part of the intestinal microbiota of healthy individuals, as well as on their active molecules; Furthermore, their selection for the development of applications should be based not on mere associations drawn from observational studies, but on direct evaluations of their efficacy in models of these pathologies. These studies can offer attractive strategies for the management of mood diseases that allow reducing their economic and social impact.
    [0021] DETAILED DESCRIPTION OF THE INVENTION
    [0023] The present invention relates to the Christensenella minuta strain DSM 32891 (C. minuta DSM 32891), to the cellular components, metabolites, secreted molecules of said strain, and to the compositions comprising the above products, as well as to their use for the prevention and / or treatment of mood disorders, such as depression.
    [0025] The inventors have discovered that strain C. minuta DSM 32891 exhibits the ability to attenuate depressive behavior in animals exposed to acute social stress. This effect has been demonstrated by orally administering the bacteria ( C. minuta strain DSM 32891) to an animal model of social stress that induces depressive symptoms (see Example 2).
    [0027] Therefore, in one aspect, the present invention relates to Christensenella minuta strain DSM 32891, hereinafter "strain of the invention", "C. minuta strain DSM 32891" or "strain DSM 32891".
    [0029] The C. minuta DSM 32891 strain was isolated from feces from healthy humans. The strain was deposited by the applicant on August 7, 2018 under the Treaty of Budapest in the German Type Culture Collection (DSM) as the International Depository Authority Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Cultures, InhoffenstraBe 7B, 38124 Braunschweig, GERMANY). The assigned deposit number was DSM 32891.
    [0031] The scientific classification of the strain of the invention is: Domain: Bacteria; Edge: Firmicutes; Class: Clostridia; Order: Clostridiales; Family: Christensenellaceae; Species: C. minuta.
    [0033] Another aspect of the present invention relates to a strain derived from the C. minuta DSM 32891 strain, wherein said strain maintains or improves the capabilities described throughout the present invention. The derived microorganism can be produced naturally or intentionally, by mutagenesis methods known in the state of the art, such as, but not limited to, the growth of the original microorganism in the presence of mutagenic or stress-causing agents, or by engineering. genetics aimed at modifying specific genes. According to a preferred embodiment, the strain derived from the C. minuta DSM 32891 strain is a genetically modified mutant. The terms mutant strain or derived strain can be used interchangeably.
    [0035] The C. minuta DSM 32891 strain or any mutant or derivative thereof can be used in any way that exerts the described effects, such as, according to a preferred embodiment of the present invention, the C. minuta DSM 32891 strain is in the form of viable cells (cultivable or non-cultivable), or according to another preferred embodiment of the invention the strain is in the form of non-viable cells ("dead" cells inactivated by any technique known in the state of the art such as, but not limited to , heat, freezing or ultraviolet radiation).
    [0037] Another aspect of the present invention refers to the cellular components, metabolites, secreted molecules or any of their combinations, obtained from the strain of the invention, or from a combination of microorganisms comprising at least one strain of the invention.
    [0039] The cellular components of the bacterium could include components of the cell wall (such as but not limited to, peptidoglycan), nucleic acids, membrane components, or others such as proteins, lipids, and carbohydrates and their combinations, such as lipoproteins, glycolipids or glycoproteins. Metabolites include any molecule produced or modified by the bacterium as a consequence of its metabolic activity during its growth, its use in technological processes (for example, but not limited to, food or drug manufacturing processes), during product storage or during gastrointestinal transit. Examples of these metabolites are, but are not limited to, organic and inorganic acids, proteins, peptides, amino acids, enzymes, lipids, carbohydrates, lipoproteins, glycolipids, glycoproteins, vitamins, salts, metals, or nucleic acids. Secreted molecules include any molecule exported or released abroad by the bacterium during its growth, its use in technological processes (for example, food or drug manufacturing), product storage, or gastrointestinal transit. Examples of these molecules are, but are not limited to, organic and inorganic acids, proteins, peptides, amino acids, enzymes, lipids, carbohydrates, lipoproteins, glycolipids, glycoproteins, vitamins, salts, metals, or nucleic acids.
    [0041] Another aspect of the present invention relates to a composition, hereinafter "composition of the invention", comprising the strain of the invention and / or the cellular components, metabolites, secreted molecules of the strain of the invention or any of their combinations.
    [0043] The composition, defined generally, is a set of components that is formed by at least the strain of the invention in any concentration; or at least by the cellular components, metabolites, secreted molecules of the strain of the invention or any of its combinations; or by a combination thereof.
    [0045] In a preferred embodiment, the composition of the invention has a concentration of the strain of the invention of between 104 and 1014 colony forming units (cfu) per gram or milliliter of final composition.
    [0047] In another particular embodiment, the composition of the invention may further comprise at least one additional microorganism other than the strain of the invention and / or its cellular components, metabolites or secreted molecules, or any combination thereof. For example, but not limited to, the additional microorganism that may be part of said composition is selected from at least one of the following groups:
    [0048] - at least one strain of another species of the genus Chrístensenella and, especially, or of the species Chrístensenella minuta;
    [0049] - at least one lactic bacteria or bifidobacterium of intestinal, food or environmental origin. The lactic acid bacterium is selected from the list comprising, but not limited to, a bacterium of the genus Bifidobacterium, Lactobacillus, Lactococcus, Enterococcus, Propionibacterium, Leuconostoc, Weissella, Pediococcus, or Streptococcus;
    [0050] - at least one strain of other phylogenetic groups, genera or species of prokaryotes of intestinal, food or environmental origin, such as but not limited to Archaea, Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria, Verrucomicrobia, Fusobacteria, Metanobacteria, Spirochaetes, Fibrobacteres, Deferribacteres, Deinococcus, Thermus, Cyanobacteria, Methanobrevibacterium, Peptostreptococcus, Ruminococcus, Coprococcus, Subdolingranulum, Dorea, Bulleidia, Anaerofustis, Bacteria, Catenibacterium, Dialister, Anaerotruncus, Anaerotruncus Eubacterium, Akkermansia, Bacillus, Butyrivibrio or Clostridium;
    [0051] - at least one strain of fungus or yeast such as, but not limited to, belonging to the genus Saccharomyces, Candida, Pichia, Debaryomyces, Torulopsis, Aspergillus, Rhizopus, Mucor or Penicillium .
    [0053] Said additional microorganism may be a strain of the same species or of a different species or taxonomic group of microorganisms from that corresponding to the strain of the invention. The cells that comprise the composition can be non-viable or viable and be in any stage of the state of development or growth (latent, exponential, stationary, etc.), regardless of the morphology it presents. In a particular embodiment, said additional microorganism comprises at least one intestinal bacterium or a lactic acid bacterium.
    [0055] Optionally, in another particular embodiment, the composition of the invention may further comprise at least one bioactive component (active substance, active ingredient or therapeutic agent), such as for example components of food, plant products and / or drugs.
    [0057] The term "bioactive component" refers to a compound with activity biological in the scope of the patent that can improve or complement the activity of the C. minuta DSM 32891 strain, including ingredients or components of food (for example and without limitation: polyunsaturated fatty acids, conjugated linoleic acid, prebiotics , fiber, Guar gum, glucomannan, chitosan, copper picolinate, calcium, etc.), other probiotics, plants, extracts or components of plants and drugs.
    [0059] In a particular embodiment, the composition of the invention is a pharmaceutical composition. The pharmaceutical composition is a set of components that is formed by at least the strain of the invention in any concentration; or at least by the cellular components, metabolites, secreted molecules of the strain of the invention or any of their combinations, which has at least one application in the improvement of the physical or physiological or psychological well-being of a subject, which implies an improvement of the state general health or reduced risk of disease. Said pharmaceutical composition may be a medicine.
    [0061] The term "medicine" has a more limited meaning than the meaning of "pharmaceutical composition", as defined in the present invention, since the medicine necessarily implies a preventive or therapeutic effect. The medicine to which the present invention refers can be for human or veterinary use. The "medicine for human use" is any substance or combination of substances that is presented as possessing properties for the treatment or prevention of diseases in humans or that can be used in humans or administered to humans in order to restore, correct or to modify the physiological functions exerting a pharmacological, immunological or metabolic action, or to establish a medical diagnosis. "Veterinary medicine" is any substance or combination of substances that is presented as having curative or preventive properties with respect to animal diseases or that can be administered to the animal in order to restore, correct or modify its physiological functions by exercising a pharmacological, immunological or metabolic action, or to establish a veterinary diagnosis. "Veterinary drugs" are also considered "veterinary feed premixes" prepared for incorporation in feed.
    [0063] In addition to the requirement of therapeutic efficacy where said composition pharmaceutical may require the use of other therapeutic agents, there may be additional fundamental reasons that strongly compel or recommend the use of a combination of a compound of the invention and a biactive component, where said bioactive component is attributed an appropriate activity for constitute a medicine. Said compound of the invention obviously refers to the strain of the invention, or to the strain derived from it, or to the cellular components, metabolites, secreted molecules or any of their combinations, obtained from the strain of the invention.
    [0065] In a particular embodiment, the pharmaceutical composition further comprises at least one pharmaceutically acceptable carrier and / or excipient.
    [0067] The "vehicle" or carrier is preferably an inert substance. The function of the vehicle is to facilitate the incorporation of other compounds, to allow for better dosing and administration, or to give consistency and shape to the pharmaceutical composition. Therefore, the vehicle is a substance which it is used in the medicine to dilute any of the components of the pharmaceutical composition of the present invention up to a determined volume or weight, or even without diluting said components it is capable of allowing better dosage and administration or giving consistency and shape to the medicine When the presentation form is liquid, the pharmaceutically acceptable vehicle is the diluent.
    [0069] The term "excipient" refers to a substance that aids in the absorption of any of the components of the composition of the present invention, stabilizes said components or aids in the preparation of the pharmaceutical composition in the sense of giving it consistency or providing flavors that Thus, the excipients could have the function of keeping the components together, such as starches, sugars or cellulose, sweetening function, dye function, protection function of the medicine, for example, to isolate it from the air and / or or humidity, the filling function of a pill, capsule or any other form of presentation such as, for example, dibasic calcium phosphate, a disintegrating function to facilitate the dissolution of the components and their absorption in the intestine, without excluding other types of excipients not mentioned in this paragraph. Therefore, the term "excipient" is defined as that matter that, included in the forms ga lenicas, is added to the active substances or their associations to enable its preparation and stability, modify its organoleptic properties or determine the physico-chemical properties of the pharmaceutical composition and its bioavailability. The "pharmaceutically acceptable" excipient must allow the activity of the compounds of the pharmaceutical composition, that is, to be compatible with said components.
    [0071] Furthermore, as understood by the person skilled in the art, the excipient and the vehicle must be pharmacologically acceptable, that is, that the excipient and the vehicle are allowed and evaluated so as not to cause harm to the organisms to which it is administered.
    [0073] The pharmaceutical composition or medicament can be presented under any clinically permitted administration form and in a therapeutically effective amount. For example, a form adapted to oral, sublingual, nasal, intracathecal, bronchial, lymphatic, rectal, transdermal, inhaled, or parenteral administration may be in form, preferably in a form adapted to oral administration. The pharmaceutical composition of the invention can be formulated in solid, semi-solid, liquid or gaseous forms, such as tablet, capsule, powder, granule, ointment, solution, suppository, injection, inhalant, gel, microsphere or aerosol. The form adapted for oral administration is selected from the list comprising, but not limited to, drops, syrup, herbal tea, elixir, suspension, extemporaneous suspension, drinkable vial, tablet, capsule, granule, seal, pill, tablet, pill, troche or lyophilized. In a particular embodiment, the composition of the invention is presented in a composition adapted to oral, sublingual, nasal, bronchial, lymphatic, rectal, transdermal, inhaled or parenteral administration.
    [0075] In a more particular embodiment, the composition of the invention is presented in a form adapted for oral administration. The form adapted for oral administration refers to a physical state that can allow oral administration. Said form adapted for oral administration is selected from the list comprising, but not limited to, drops, syrup, herbal tea, elixir, suspension, extemporaneous suspension, drinkable vial, tablet, capsule, granule, seal, pill, tablet, pill, troche or lyophilized.
    [0077] The "pharmaceutical form" or "pharmaceutical form" is the disposition to which the active ingredients and excipients are adapted to constitute a medicine. It is defined by the combination of the way in which the pharmaceutical composition is presented by the manufacturer and the way in which it is administered.
    [0079] In the present invention, the term "therapeutically effective amount" refers to that amount of the component of the pharmaceutical composition that when administered to a mammal, preferably a human, is sufficient to produce prevention and / or treatment, such as defined below, of a disease or pathological condition of interest in the mammal, preferably a human The therapeutically effective amount will vary, for example, according to the activity of the strain of the invention, of the cellular components, metabolites, secreted molecules or any of its combinations, in any form of presentation; the therapeutically effective amount will also vary according to the metabolic stability and duration of action of the compound; the patient's age, body weight, general health, sex and diet ; mode and time of administration; rate of excretion, drug combination; severity of disorder or condition p particular atological; and the subject undergoing therapy, but may be determined by a specialist in the art based on his or her own knowledge and description.
    [0081] As an alternative to the pharmaceutical composition, the composition of the invention can also be a nutritional composition.
    [0083] The term "nutritional composition" of the present invention refers to that food that, independently of providing nutrients to the subject who takes it, beneficially affects one or more functions of the organism, so as to provide a better state of health and well-being. As a consequence, said nutritional composition can be intended for the prevention and / or treatment of a disease or the factor causing a disease. Therefore, the term "nutritional composition" of the present invention can be used as a synonym for functional food or food for specific nutritional purposes or medicinal food.
    [0085] In a particular embodiment, the nutritional composition is a food, a supplement, a nutraceutical, a probiotic, or a symbiotic.
    [0087] In a more particular embodiment, the food is selected from the list comprising: dairy product, vegetable product, meat product, snack, chocolate, drink or infant food. The dairy product is selected from the list which comprises, but is not limited to, product derived from fermented (eg, but not limited to yogurt or cheese) or unfermented (eg, but not limited to) ice cream, butter, margarine, dairy whey ). The plant product is, for example, but not limited to, a cereal in any form, fermented or unfermented. The beverage can be, but is not limited to, any fruit juice or unfermented milk.
    [0089] The term "supplement", synonymous with any of the terms "dietary supplement", "nutritional supplements"; or "food supplement" is a "food ingredient" intended to supplement food. Some examples of dietary supplements include, but are not limited to, vitamins, minerals, botanicals, amino acids, and food components such as enzymes and glandular extracts. They are not presented as substitutes for a conventional food or as a single component of a meal or diet but rather as a supplement to the diet.
    [0091] The term "nutraceutical" as used in the present invention refers to substances isolated from a food and used in dosages that have a beneficial effect on health.
    [0093] The term "probiotic" as used in the present invention refers to live microorganisms that when supplied in adequate amounts promote health benefits of the host organism.
    [0095] The term "symbiotic" as used in the present invention refers to those foods that contain a mixture of prebiotics and probiotics. As a general rule, they contain a prebiotic component that favors growth and / or metabolic activity and, in short, the effect of the probiotic with which it is combined, as for example and without limitation, may be the association of fructooligosaccharides or galactooligosaccharides with bifidobacteria.
    [0097] Another aspect of the present invention relates to the use of the strain of the invention, or the components derived from it, or the composition of the invention, for the manufacture of a medicine, a nutritional composition or a food.
    [0099] Another aspect of the present invention relates to strain C. minuta DSM 32891, a cellular component, metabolite, secreted molecule or any of its combinations obtained from the strain of the invention, or the composition of the invention, for use as a medicine. The term medicament has been previously defined, and is applicable to the present inventive aspect.
    [0101] In another aspect, the present invention relates to the strain of the invention, a strain derived from it, a cellular component, metabolite, secreted molecule or any of its combinations obtained from the strain of the invention, or the composition of the invention, for use in the prevention and / or treatment of mood disorders, such as depression.
    [0103] In the present invention, the term "treatment" refers to combating the effects caused as a consequence of a disease or pathological condition of interest in a subject (preferably mammal, and more preferably a human) including: (i) inhibiting the disease or pathological condition, that is, stopping its development;
    [0104] (ii) alleviating the disease or the pathological condition, that is, causing the regression of the disease or the pathological condition or its symptoms;
    [0105] (iii) stabilize the disease or pathological condition.
    [0107] In the present invention, the term "prevention" refers to preventing the occurrence of the disease, that is, preventing the disease or pathological condition from occurring in a subject (preferably a mammal, and more preferably a human), in particular, when said subject has a predisposition for the pathological condition.
    [0109] In the present invention, mood disorders or disturbances include, but are not limited to, depression, major depression, atypical depression, typical or melancholic depression, psychotic depression, catatonic depression, pre- and postpartum depression, bipolar disorder, seasonal affective disorder , dysthymia, depressive personality disorder, double depression, unspecified depressive disorder, recurrent brief depressive disorder, minor depression, mood disorder induced by substance abuse or drug use, etc.
    [0111] In another aspect, the present invention relates to the strain of the invention, the strain derived from the strain of the invention, the cellular component, metabolite, secreted molecule or any of its combinations obtained from the strain of invention, or the composition of the invention, for use as an adjunct in the treatment or prevention of mood disorders.
    [0113] In the present invention, "adjuvant" is understood to be that compound that helps to improve the effectiveness or efficiency of other medications for the treatment of mood disorders, which would allow to reduce their dose and / or the frequency of administration or enhance its efficacy by administering a formulation of the strain of the invention with complementary mechanisms of action.
    [0115] In another aspect, the present invention relates to the use of the strain of the invention, a strain derived from the strain of the invention, the cellular component, metabolite, secreted molecule or any of its combinations obtained from the strain of invention, or the composition of the invention, for the elaboration of a food. The term medicament has been previously defined in the present description and is applicable to the present inventive aspect.
    [0117] Throughout the description and claims, the word "comprises" and its variants are not intended to exclude other technical characteristics, additives, components or steps. For those skilled in the art, other objects, advantages, and features of the invention will emerge in part from the description and in part from the practice of the invention. The following examples and figures are provided by way of illustration, and are not intended to be limiting of the present invention.
    [0119] BRIEF DESCRIPTION OF THE FIGURES
    [0121] Fig. 1: Evaluation of the effect of the administration of B. breve MF217 (1x109 cfu / day) and C. minuta DSM 32891 (1x109 cfu / day) in C57BL / 6 mice (n = 10 / group) exposed to acute social stress in the sucrose preference test (SPF). Data are expressed in grams with means and standard error. Statistically significant differences were established by applying a one-way ANOVA followed by the Bonferroni post hoc test (*** = p <0.001, different from control mice; # = p <0.05, different from mice subjected to stress and without try). C, control mice; E, mice subjected to untreated acute stress; B. Bre, mice under acute stress and treated with B. breve; C. min, mice under acute stress and treated with C. minuta.
    [0122] Fig. 2: Evaluation of the effect of the administration of strain B. breve MF217 (1x109 cfu / day) and C. minuta DSM 32891 (1x109 cfu / day) in C57BL / 6 mice (n = 10 / group) exposed to stress Social Acute in the Tail Suspension Test (TST). Data are expressed as means of time (seconds) and their standard error. Statistically significant differences were established by applying a one-way ANOVA followed by the Bonferroni post hoc test (*** = p <0.001, different from control mice). C, control mice; E, mice subjected to untreated acute stress; B. Bre, mice under acute stress and treated with B. breve; C. Min, mice under acute stress and treated with C. minuta.
    [0124] EXAMPLES
    [0126] The invention will now be illustrated by tests carried out by the inventors, which shows the effectiveness of the product of the invention.
    [0128] EXAMPLE 1. Isolation and identification of Christensenella species
    [0129] The biological material object of the patent was isolated from faeces from healthy volunteers, which were processed and inoculated in Medium for Intestinal Bacteria (MBI) whose composition is based on the means recommended in previous publications [18], [19], modifications designed by the inventors. These consisted of fermenting the processed feces, maintaining a constant pH, in an anaerobic chamber, for 24 hours. The fermented MBI medium itself was used as a supplement to the Fastidious Anaerobe Agar (FAA) medium with 0.5% defibrillated blood, which was used to inoculate serial dilutions of the faeces and isolate colonies, after incubating the plates for 72h at 37 ° C and in an anaerobiosis chamber. Among the growing colonies, Christensenella minuta DSM 32891 was isolated.
    [0131] Its identification was performed by sequencing the 16S rRNA gene (1.26 Kb) using primers 27f (SEQ ID NO: 1: 5'-AGAGTTTGATCCTGGCTCAG-3 ') and 1401r (SEQ ID NO: 2: 5'- CGGTGTGTACAAGACCC-3' ) by Sanger technology on an ABI 3730XL sequencer. By comparing the sequences obtained with those of the NCBI database and the BLASTn algorithm, the identification of the isolated strain DSM 32891 with the species Christensenella minuta was obtained with a percentage 100% identity. The complete sequence (SEQ ID NO: 3) is included below:
    [0133] > 16S rRNA gene sequence of Christensenella minuta DSM 32891
    [0134] ACTTCATGTGGGCGGGTTGCAGCCCACAAT CCGAACTGGGACCGGCTTTTTGAGATTCGC TTCCCCTTACGGGTTCGCTGCCCTTTGTACCGGCCATTGTAGCACGTGTGTAGCCCAAGA CATAAGGGGCATGATGATTTGACGTCGTCCCCACCTTCCTCCGAGTTGTCCCCGGCAGTC TCACTAGAGTTCCCGCCTTTACGCGCTGGCAACTAGCAATAAGGGTTGCGCTCGTTGCGG GACTTAACCCAACATCTCACGACACGAGCTGACGACAACCATGCACCACCTGTCTCTCTG CCCCGAAGGGAAACTGTATCTCTACAGTCGTCAGAGGATGTCAAGCCTTGGTAAGGTTCT TCGCGTTGCTTCGAATTAAACCACATGCTCCGCTGCTTGTGCGGGCCCCCGTCAATTCCT TTGAGTTTCAACCTTGCGATCGTACTCCCCAGGCGGGATACTTAATGCGTTTGCTTCGGC ACGGAACCCTATCGGGCCCCACACCTAGTATCCATCGTTTACGGCGTGGACTACCAGGGT ATCTAATCCTGTTTGCTCCCCACGCTTTCGTGCCTCAGTGTCAGTTACAGTCCAGAAAGT CGCCTTCGCCACTGGTGTTCCTCCTAATATCTACGCATTTCACCGCTACACTAGGAATTC CACTTCCCTCTCCTGTACTCAAGTCACACAGTTTCAAATGCAACCCCGGGGTTAAGCCCC GGTCTTTCACATCTGACTTACATGACCACCTACGCACCCTTTACGCCCAGTAATTCCGGA CAACGCTTGCTCCCTACGTATTACCGCGGCTGCTGGCACGTAGTTAGCCGGAGCTTCCTC CTATGGTACCGTCATTTCTTTCGTCCCATAGGACAAAGGTTTACAATCCGAAGACCTTCT TCCCTCACGCGGCGTTGCTGGGTCAGGGTTTCCCCCATTGCCCAATATTCCCCACTGCTG CCTCCCGTAGGAGTCTGGACCGT GTCTCAGTTCCAGTGTGGCCGATCACCCTCTCAGGTC GGCTACCCATCGTTGACTTGGTGGGCCGTTACCTCACCAACTATCTAATGGGACGCGAGC CCATCCTGCATCGAATAAATCCTTTTACCTCAAAACCATGCGGTTTCGTGGTCTCATGCG GTATTAGCAGTCGTTTCCAACTGTTGTCCCCCGTTGCAGGGCAGGTTGCTCACGCGTTAC TCACCCGTCCGCCACTCGGTATACCCACAGTTCCTCCCGAAGGATTCACAAAGGGCAACC T
    [0135] EXAMPLE 2. Effects of C. minuta strain DSM 32891 in an animal model of depression induced by social stress.
    [0136] Development of the animal model of acute social stress and sampling
    [0137] Male C57BL / 6 mice that came of adolescent age (postnatal day 32, Charles River, Les Oncins, France) were used for this study. They were kept under controlled conditions of temperature (23 ° C), relative humidity (40-50%) and a light / dark cycle of 12 hours and fed a standard diet (D12450K, Research diet, Brogaarden, Denmark) for 11 weeks (the first week they were quarantined in a room prepared to prevent possible zoonoses). Mice were randomly divided into 4 experimental groups (n = 10). Control group (C), stress group (E), group treated with Christensenella minuta DSM 32891 (C.min) and group treated with Bifidobacterium breve MF217 (B. breve) used for comparative purposes. Mice in the C. min group were treated with an oral dose of the bacterial strain object of the invention (1x109 colony forming units [CFU]) suspended in 10% skimmed milk; group B. brief was treated with one dose B. brief (1x109 colony forming units [CFU]) suspended in 10% skim milk. The vehicle or placebo (10% skimmed milk) was administered in the same way to both the control group and the stress group. Treatment or placebo was administered for 10 weeks. At the end of these 10 weeks, the mice were sacrificed by cervical dislocation to obtain samples, including blood, intestine, brain, fecal content and feces.
    [0139] Acute social stress model
    [0140] To induce acute social stress, an animal model of social defeat based on the resident-intruder paradigm was used [20]. In this model, one of the two animals (the resident) was allowed to establish territoriality in its own box. Next, the mice under study (intruder), in our case C57BL / 6 males, were introduced one by one in the cage of the resident mouse. For this, aggressive adult males (4 weeks) of the CD-1 strain (Charles River, Les Oncins, France), who were previously isolated and trained to be more aggressive, were used as resident mice. Agonistic encounters were carried out for 4 consecutive days (introduction of the naive mouse into the resident's cage for 10 minutes) in which physical contact was allowed between them and in which the intruder mouse suffered a high degree of stress (reflected in the production of high levels of corticosterone). The agonistic encounters took place in a neutral room and not in the animal house where they were usually housed. The experimental mice (intruders) exhibited avoidance or flight behavior, as well as defense / submission behavior after suffering aggression (threat / attack) from their opponent. The criterion used to define that an animal had been defeated was the adoption of a specific posture that means defeat. It is characterized by a posture of vertical submission with flaccid front legs, head tilted upwards and ears retracted [21].
    [0142] The control group was not exposed to social defeat; however, all the mice in this group were placed 10 minutes in a cage exactly the same as those used to perform the agonistic encounters. For 10 minutes they explored the cage without having contact with any opponent.
    [0144] Before performing the agonistic encounters, the animals fasted for 12 hours. Immediately after the social defeat the animals were exposed for 2 hours to food, water and water with 3% sucrose.
    [0146] Sucrose Preference Test 3% Sucrose Preference Test (SPT)
    [0147] The 3% sucrose preference test was performed to evaluate the hedonic / anhedonic behavior associated with depressive behavior. Anhedonic behavior (inability to feel pleasure) is considered one of the clearest symptoms of depression [22]. Different animal studies have shown that depressed animals consume less water with 3% sucrose, considering this an anhedonic behavior.
    [0149] The test consists of depriving the animals of water for 12 hours and then exposing them to two options, either water or water with 3% dissolved sucrose. The sucrose and water bottles were changed during the 2 h trial period to ensure that there was no effect related to location preference. The amount of 3% sucrose ingested during those 2h would indicate hedonic / anhedonic behavior. A lower intake of sucrose would indicate anhedonia. Sucrose preference was calculated as the percentage of sucrose ingested in relation to the total amount of liquid consumed and corrected for body weight.
    [0151] The results (Figure 1) indicate that stressed animals (E) ingest significantly less 3% sucrose than control mice (p <0.01), indicating anhedonic and, therefore, depressive behavior. Treatment with B. breve partially restores anhedonia, although this improvement is not significant. However, treatment with C. minuta does totally restore depressive behavior (p <0.05), indicating greater effectiveness compared to B. brief.
    [0153] Tail suspension test
    [0154] The mice were suspended on the edge of a table with duct tape placed approximately 1 cm from the tip of the tail in a position that they could not escape or cling to nearby surfaces. The behavior aimed at trying to escape was quantified, as well as the immobility time for 5 minutes. The duration of immobility (as a measure of demotivation) was recorded during the 5 min that the test lasted. This test is widely used for the evaluation of depressive behavior in mice [21].
    [0155] The results (Figure 2) indicate that the stressed animals (E) remain significantly longer immobile with respect to the time they are moving (p <0.001) while the control mice (C) show no significant difference. This indicates depressive behavior since the animals do not try to escape, but rather give up showing little motivation for survival. Treatment with B. breve did not improve this depressive behavior, maintaining significant differences between the time of immobility and the time that the mice were moving (p <0.001). However, treatment with C. minuta (C. min) does restore this depressive behavior, reducing the time in which the mice are immobile, so that the differences between both measures were reduced and were not significant.
    [0157] These results demonstrate that oral treatment with C. minuta shows greater efficacy compared to possible conventional probiotics, such as B. breve, in improving depressive behavior in a model of depression induced by acute social stress.
    [0159] The results of both tests show for the first time that treatment with C. minuta restores depressive behavior in mice that have been exposed to acute social stress. The data shows that C. minuta would be a better choice as a treatment to improve mood disorders, such as depressive behavior, than conventional probiotics.
    [0161] Bibliography
    [0162] 1. Gustavsson, A., et al., Cost of disorders of the brain in Europe 2010. Eur Neuropsychopharmacol, 2011. 21 (10): p. 718-79.
    [0163] 2. Lamers, F., et al., Evidence for a differential role of HPA-axis function, inflammation and metabolic syndrome in melancholic versus atypical depression. Mol Psychiatry, 2013. 18 (6): p.692-9.
    [0164] 3. Milaneschi Y, LF, Bot M, Drent ML, Penninx BW, Leptin Dysregulation Is Specifically Associated With Major Depression With Atypical Features: Evidence for a Mechanism Connecting Obesity and Depression. . Biol Psychiatry. , 2017. May 1; 81 (9): p. 807-814.
    [0165] 4. Zhernakova, A., et al., Population-based metagenomics analysis reveals markers for gut microbiome composition and diversity. Science, 2016.
    [0166] 352 (6285): p. 565-9.
    [0167] 5. McDonald, D., et al., American Gut: an Open Platform for Citizen Science Microbiome Research. mSystems, 2018. 3 (3).
    [0168] 6. Aizawa, E., et al., Possible association of Bifidobacterium and Lactobacillus in the gut microbiota of patients with major depressive disorder. J Affect Disord, 2016. 202: p.254-7.
    [0169] Jiang, H., et al., Altered fecal microbiota composition in patients with major depressive disorder. Brain Behav Immun, 2015. 48: p. 186-94.
    [0170] Kelly, JR, et al., Transferring the blues: Depression-associated gut microbiota induces neurobehavioural changes in the rat. J Psychiatr Res, 2016. 82: p. 109 18.
    [0171] Naseribafrouei A, HK, Avershina E, Sekelja M, Linl0kken A, Wilson R, Rudi K., Correlation between the human fecal microbiota and depression. Neurogastroenterol Motil. , 2014. Aug; 26 (8): p. 1155-62.
    [0172] Gibson-Smith, D., et al., The role of obesity measures in the development and persistence of major depressive disorder. J Affect Disord, 2016. 198: p.222-9. De Palma, G., et al., Microbiota and host determinants of behavioral phenotype in maternally separated mice. Nat Commun, 2015. 6: p. 7735.
    [0173] Sanz, Y., Bifidobacteria in foods: health effects, in The Encyclopedia of Food and Health. Caballero, P. Finglas, and F. Toldrá, Editors. Oxford Academic Press: Oxford., 2016: pp 388-394.
    [0174] Dinan, TG and JF Cryan, Mood by microbe: towards clinical translation. Genome Med, 2016. 8 (1): p. 36.
    [0175] Romijn, AR, et al., A double-blind, randomized, placebo-controlled trial of Lactobacillus helveticus and Bifidobacterium longum for the symptoms of depression. Aust NZJ Psychiatry, 2017. 51 (8): p. 810-821.
    [0176] Pinto-Sanchez, MI, et al., Probiotic Bifidobacterium longum NCC3001 Reduces Depression Scores and Alters Brain Activity: A Pilot Study in Patients With Irritable Bowel Syndrome. Gastroenterology, 2017. 153 (2): p.448-459 e8. Yu, M., et al., Variations in gut microbiota and fecal metabolic phenotype associated with depression by 16S rRNA gene sequencing and LC / MS-based metabolomics. J Pharm Biomed Anal, 2017. 138: p.231-239.
    [0177] I. Mironova, IZ, N. Zhukova, V. Alifirova, A. Latypova, O. Izhboldina, M. Nikitina, V. Petrov, M. Titova, Affective disorders and microbiome in patients with Parkinson's disease ( PR2063). European Journal of Neurology., 2017. 24 (Suppl. 1) ,: p.445-678.
    [0178] Gibson GR, CJ, Macfarlane GT., Use of a three-stage continuous culture system to study the effect of mucin on dissimilatory sulfate reduction and methanogenesis by mixed populations of human gut bacteria. Appl Environ Microbiol. 1988 Nov; 54 (11), 1988. Nov; 54 (11): p.2750-5.
    [0179] Lesmes, U., et al., Effects of resistant starch type III polymorphs on human colon microbiota and short chain fatty acids in human gut models. J Agric Food Chem, 2008. 56 (13): p. 5415-21.
    [0180] Tamashiro, KL, MM Nguyen, and RR Sakai, Social stress: from rodents to primates. Front Neuroendocrinol, 2005. 26 (1): p.27-40.
    [0181] Can, A., et al., The tail suspension test. J Vis Exp, 2012 (59): p. e3769.
    [0182] Duncko, R., et al., Altered function of peripheral organ systems in rats exposed to chronic mild stress model of depression. Cell Mol Neurobiol, 2001. 21 (4): p.
    [0183] 403-11.
    权利要求:
    Claims (21)
    [1]
    1. A strain of Chrístensenella minuta with deposit number DSM 32891.
    [2]
    2. A strain derived from the strain according to claim 1.
    [3]
    3. Strain according to claim 1 or 2, wherein the strain is a genetically modified mutant.
    [4]
    4. Strain according to any of claims 1 to 3, wherein said strain is in the form of viable cells or in the form of non-viable cells.
    [5]
    5. A cellular component, metabolite, secreted molecule or any of its combinations obtained from the strain according to any one of claims 1 to 4.
    [6]
    6. A composition comprising a strain according to any of claims 1 to 4 or a cellular component, metabolite, secreted molecule or any of its combinations according to claim 5.
    [7]
    7. Composition according to claim 6, wherein the composition additionally comprises at least one bioactive component.
    [8]
    8. Composition according to claim 6 or 7, wherein the composition additionally comprises at least one microorganism other than the strain according to any one of claims 1 to 4.
    [9]
    9. Composition according to claim 8, wherein the microorganism is an intestinal bacterium or a lactic bacteria.
    [10]
    Composition according to any one of Claims 6 to 9, wherein said composition is a pharmaceutical composition.
    [11]
    11. Composition according to claim 10, wherein the composition additionally comprises at least one vehicle and / or one pharmaceutically acceptable excipient.
    [12]
    12. Composition according to any one of claims 10 or 11, wherein said composition is presented in a form adapted for oral, sublingual, nasal, bronchial, lymphatic, rectal, transdermal, inhaled or parenteral administration.
    [13]
    13. Composition according to any one of Claims 6 to 9, wherein said composition is a nutritional composition.
    [14]
    14. Composition according to claim 13, wherein the nutritional composition is a food, a supplement, a nutraceutical, a probiotic or a symbiotic.
    [15]
    15. Composition according to claim 14, wherein said food is selected from the list consisting of a dairy product, a vegetable product, a meat product, a snack, chocolate, drink or baby food.
    [16]
    16. Composition according to any one of Claims 6 to 15, wherein said composition has a concentration of the strain of between 104 and 1014 colony forming units (cfu) per gram or milliliter of final composition.
    [17]
    17. Strain according to any one of claims 1 to 5, a cellular component, metabolite, secreted molecule or any of its combinations according to claim 5, or a composition according to any one of claims 6 to 16, for use as a medicine.
    [18]
    18. Strain according to any one of claims 1 to 5, a cellular component, metabolite, secreted molecule or any of its combinations according to claim 5, or a composition according to any one of claims 6 to 16, for use in prevention. and / or the treatment of mood disorders.
    [19]
    19. Strain according to any one of claims 1 to 5, a cellular component, metabolite, secreted molecule or any of its combinations according to claim 5, or a composition according to any one of claims 6 to 16, for use as an adjuvant in treatments for mood disorders.
    [20]
    twenty.
    [21]
    21. Use of the strain according to any one of claims 1 to 5, a cellular component, metabolite, secreted molecule or any of its combinations according to claim 5, or a composition according to any one of claims 6 to 16, for the preparation of a food.
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    EP3887504A1|2021-10-06|
    IL283418D0|2021-07-29|
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    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    WO2004098622A2|2003-05-08|2004-11-18|Alimentary Health Limited|Probiotics in the treatment of atypical depression and other disorders characterized by hypothalamic-pituitary-adrenal axis over-activity|
    EP2937424A1|2014-04-23|2015-10-28|National Yang-Ming University|Lactic acid bacterium, composition containing the same and their use|
    US20170042948A1|2014-04-23|2017-02-16|Cornell University|Modulation of fat storage in a subject by altering population levels of christensenellaceae in the gi tract|
    US20180255819A1|2017-03-10|2018-09-13|International Nutrition Research Company|Specific bacteria for their use as a medicament, in particular for controlling excess weight, obesity, cardiometabolic diseases and inflammatory bowel diseases|
    WO2017160711A1|2016-03-14|2017-09-21|Holobiome, Inc.|Modulation of the gut microbiome to treat mental disorders or diseases of the central nervous system|
    WO2018002238A1|2016-07-01|2018-01-04|Nestec S.A.|Nutritional composition comprising a probiotic for the prevention and/or treatment of anxiety disorders and related conditions in a mammal|
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