Method, comprising bacteriophages, for reducing population of at least one adipogenic bacterial species, and bacteriophages and the use thereof

Abstract

To provide a method for reducing the population of at least one adipogenic bacterial species.SOLUTION: A method for reducing the population of at least one adipogenic bacteria species is provided which comprises the following steps: a) providing a biologic sample which comprises at least one adipogenic bacteria species; b) providing at least one bacteriophage species which is specific to at least one adipogenic bacteria species and comprises at least one nucleic acid functionally bound to a promoter and / or to a regulatory element; and c) causing the biologic sample to be exposed to and incubated with the bacteriophages, the incubation taking place until the population of the adipogenic bacteria species is reduced by at least 70%. The invention also relates to bacteriophages and their use.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to a method for reducing the population of at least one lipogenic bacterial species as well as to bacteriophages and their uses according to the preambles of the independent claims. [Background technology]

[0002] It is a well-known fact that interactions between bacteria and their hosts, i.e. humans and / or animals, can have both positive and negative effects. On the one hand, bacteria can be symbionts or commensals for humans or animals, and therefore function or coexist with them without harming them. On the other hand, many bacterial species are known that can attack and damage humans or animals as pathological organisms or parasites, suspected of causing and / or resulting in a series of diseases and associated sequelae (such as the appearance of overweight or excessive weight gain). This type of damage can result in the death of the organism.

[0003] To combat bacterial colonization and infection, many chemicals have become established as antibiotics and are now considered conventional treatments for humans and animals.

[0004] These types of substances are characterized by a broad spectrum of activity, since they target bacterial molecular structures or rather molecules found in many bacterial species. This allows a wide range of bacteria to be reduced or eliminated by the antibiotic. The drawback of this effect is that the aforementioned commensal or symbiotic bacteria are also killed by the use of the antibiotic, which may result in secondary damage to humans, animals and / or the environment. For example, as a result of damage to commensal and / or symbiotic bacteria, the balance of the microbiome may shift in favor of pathogenic microorganisms (e.g. bacteria and / or yeasts), an effect that may result in a condition that may last for several months, leading to serious consequences, not uncommonly.

[0005] Also, permissible interactions between chemicals used as antibiotics and physiological metabolic processes are well known. In this case, for example, certain antibiotics are known to have anabolic properties that are exploited in agriculture. However, it is considered to be a disadvantage that antibiotics used in agriculture can be taken up by humans and / or animals via the food chain, resulting in fattening effects such as growth promotion and / or hormonal development. Other forms of incompatibility of chemicals used as antibiotics have also been described. These include, among others, the appearance of allergies to these chemicals and the effects of allergic reactions resulting therefrom, in terms of allergic shock and / or the development of various (food) intolerances. Moreover, in case of failure of antibiotic treatment, repeated treatment with the same medicinal ingredient is not possible, which means that the treatment is generally switched to a different group of antibiotic active ingredients with a broader spectrum of activity, which results in even more serious imbalances of the microflora, including the appearance of multi-resistant bacteria. Indeed, certain waiting periods must be maintained in turn between two antibiotic treatments, which may lead to the spread of pathological bacteria. In this case, as in the case of early termination of antibiotic use after a presumed successful treatment, bacteria exposed to chemicals without dying may develop what are known as resistance mechanisms and resistance to antibiotics. When resistance occurs, this particular bacterium becomes immune to antibiotic treatment with the corresponding chemical. In this case, the treatment or therapy can no longer be considered successful. This can also happen when residual amounts of antibiotics or their decomposition products actually contaminate drinking water and / or the environment, resulting in exposure of bacteria in the environment to this material. In particular, it is more difficult for chemicals used as antibiotics to penetrate biofilms formed by bacteria, which are characterized by polar surfaces, especially sugars, that some known antibiotics cannot penetrate.

[0006] Overweight, adiposity and / or excessive weight gain can be referred to as a global epidemic to date, and are especially problematic in terms of physical and mental well-being. These mental and / or physical sufferings may not only result from medical consequences caused by overweight, adiposity and / or excessive weight gain, but may also be promoted due to increased risks such as metabolic, cardiovascular, hormonal balance, musculoskeletal or other organ system disorders. These include disorders or conditions such as arteriosclerosis, diabetes, joint pain, including but not limited to those in the spine, hips and knees, indigestion and cancer, such as breast cancer, intestinal cancer and pancreatic cancer. However, weight loss in the case of overweight and / or adiposity, or optimal weight stabilization, is only possible with great difficulty for many of the affected people, as modern lifestyles and / or family trends make weight loss difficult for those who suffer from overweight, adiposity and / or excessive weight gain. However, this weight loss is essential, especially for overweight people or those who suffer from severe weight gain, because overweight, adiposity and / or excessive weight gain can cause complications in the treatment of many diseases or delay or prevent successful treatment. Moreover, excessive fattening of tissues and / or organs in overweight people can disrupt the function of organs or even cause organ failure. Further diseases, which overweight generally contributes to at least the emergence or causes complications during the treatment of these diseases, affect medical fields such as oncology, immunology, gastroenterology, endocrinology, psychiatry, psychosomatic medicine, orthopedics, pediatrics, surgery, urology, etc.

[0007] Being overweight can cause or make more difficult to treat medical conditions such as metabolic syndrome, type 2 diabetes, gallbladder disease, chronic enteritis, hyperchloremic xerosis, high blood pressure, lipid metabolism disorders, dyspnea, sleep apnea, coronary heart disease, arthropathy, gout, cancer, e.g. uterine, breast, cervical, intestinal, prostate and pancreatic cancer, sex hormone disorders, decreased libido, joint and back pain, increased risk of thrombosis and embolism, increased risks during surgery and narcotics, psychosocial problems and the resulting limitations in quality of life, resulting from depression caused by low self-esteem and / or the perception of being less valued by the environment.

[0008] The appearance of overweight can also be attributed to bacteria in the intestinal microflora. It is conceivable that a reduction in the population of each of the bacteria in the intestinal microflora would prevent or at least limit the occurrence of overweight or excessive weight gain. The drawback here is that conventional antibiotic treatments not only eliminate the target bacteria that promote overweight, but also eliminate the commensal bacteria that have a positive effect on the host organism, i.e., humans and / or animals, benefiting its digestive process. This also means further increasing the circulation of antibiotics that can increase resistance. In other respects, using antibiotics to lose weight is generally not recommended, since antibiotics must be taken for a long period (e.g., for several months), which means excessive damage to the commensal microflora and the long-term occurrence of antibiotic-related side effects. Treatment consisting of antibiotics alone is not favorable for the treatment and / or therapy of overweight or excessive weight gain. Summary of the Invention

[0009] As a result of the described drawbacks of conventional treatments with chemicals used as antibiotics, and for the conventional fight against bacterial populations in general, and in particular for the treatment or therapy of overweight, adiposity and / or excess weight gain, there is a large and continuously increasing demand for modulating, combating and / or eliminating at least one adipogenic bacterial species in a simple, effective, compatible, medically safe, inexpensive, targeted and specific manner with few side effects. At the same time, it is essential that the treatment is successful in the long term. Therefore, the object of the present invention is to provide a bacteriophage and a method for reducing the population of at least one adipogenic bacterial species, characterized by high efficacy, specificity, compatibility and / or reliability, and by a simple, time-efficient and cost-effective use with few defects, in order to prevent the drawbacks of the above-mentioned state of the art.

[0010] This object is achieved in a surprisingly simple and effective manner by the methods and bacteriophages according to the teaching of the independent claims. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] According to the present invention, a method for reducing the population of at least one adipogenic bacterial species is proposed, which comprises the following steps: a) providing a biological sample containing bacteria of at least one adipogenic bacterial species; and b) providing a bacteriophage of at least one bacteriophage species specific for at least one adipogenic bacterial species and comprising at least one nucleic acid operably linked to a promoter and / or a regulatory element, said nucleic acid being selected from the group consisting of: i) a nucleic acid sequence encoding at least one antimicrobial nucleic acid molecule; and ii) a nucleic acid sequence encoding a nucleic acid molecule, said nucleic acid sequence being at least 50% identical to the nucleic acid molecule encoded by the nucleic acid sequence in i; and iii) a nucleic acid sequence encoding at least one antimicrobial polypeptide; and iv) a nucleic acid sequence encoding a polypeptide, said nucleic acid sequence being at least 50% identical to the polypeptide encoded by the nucleic acid sequence in iii; and v) nucleic acid sequences for fragments of nucleic acids, nucleic acid molecules or polypeptide-encoding fragments from i, ii, iii, iv the steps being selected from: c) exposing the biological sample to the bacteriophage and incubating it with the bacteriophage, said incubation being carried out until the population of the lipogenic bacterial species is reduced by at least 70%; Includes.

[0012] The method according to the invention is based on the fundamental idea that by using a bacteriophage specific for at least one lipogenic bacterial species, the population of this at least one lipogenic bacterial species can be reliably, quickly, easily, specifically and persistently reduced. In doing so, it has been advantageously demonstrated that other organisms, such as yeasts, bacteria, viruses, and / or cells of multicellular tissues and / or organisms are not affected and no damage, impairment and / or destruction occurs due to the simultaneous host dependency. Due to the host dependency of the bacteriophage, the method according to the invention can be repeated at any time without any resistance occurring. As is well known to those skilled in the art, mutual host dependency exists when there is no reduction of different bacterial species in a biological sample for which the bacteriophage is not specific, when the respective lipogenic bacterial species are no longer present in the biological sample. This also concerns the organs and / or tissues of the host organism, i.e. human and / or animal tissues.

[0013] Within the scope of the present invention, it has been discovered that in order to reduce the population of at least one bacterial species, it is sufficient to provide a biological sample comprising bacteria of at least one adipogenic bacterial species and to incubate this bacterial species with a given bacteriophage of at least one bacteriophage species that is specific for at least one adipogenic bacterial species.

[0014] The term "population" is well known to those skilled in the art and generally relates to the totality of all individuals of the same species present in a particular area and / or in a particular environmental area. Within the scope of the present invention, the population relates to the totality of bacteria of at least one lipogenic bacterial species. The terms "bacterium" and "bacteria" are well known to those skilled in the art as interchangeable synonyms and refer to representatives of one or more bacterial species.

[0015] The term "adipogenic bacterial species" relates to at least one bacterial species, preferably 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more bacterial species, which are well known to the person skilled in the art and which, due to their biology, have a direct or indirect effect on humans and / or animals, for example through their molecular metabolism and / or interaction with other bacterial species, yeasts and / or cells, which initiate, support and / or increase obesity, fat, overweight and / or weight gain, by structures where adipose tissue, preferably white adipose tissue, is stored and / or reduced. The direct or indirect effect on an organism is well known to the person skilled in the art. They can, for example, directly affect an organism by destroying the organism's tissues or indirectly affect metabolic or other processes in the organism, thereby in particular initiating and / or supporting weight gain and / or non-reduction of overweight. Furthermore, the skilled artisan will also understand that "adipogenesis" in the present context also means that the corresponding bacterial species may not themselves directly cause an adipogenic effect, but may, however, oppose the growth, biology and / or survival of anti-adipogenic bacterial species, thus having an adipogenic effect on the organism. Within the scope of the present invention, it is considered that the adipogenic bacterial species may be present in the gastrointestinal tract of humans and / or animals, such as in the stomach and / or intestines.

[0016] Adipogenic bacterial species include, but are not limited to, Acinetobacter, Actinobacteria, Actinomycetes, Bacillus, Bacteridos, Bartonella, Bordetella, Borreli, Brucella, Citrobacter, Campylobacter, Chlamydia, Clostridium, Corynebacter, Desulfovibrio, Ehrlichia, Enterobacter, Enterobacteriaceae, Enterococcus, Erysipelotrichia, Escherichia, Faecalibacterium, Firmicutes, Francisella, Helicobacter, Haemophilus, Klebsiella, Lactobacillus, Legionella, Leptospira, Listeria, Methanobrevibacter, Moraxella, Mycobacterium, Mycoplasma, Neisseria, Nocardia , Oscillobacter, Paeruginosa, Prevotellaceae, Propionibacterium, Proteus, Pseudomonas, Rickettsia, Ruminococci, Salmonella, Shigella, Spirillium, Spirochetes, Staphylococcus, Stenotrophomonas, Streptococcus, Treponema, Vibrio, Yersenia, Bifidobacterium, Blautia, Bradya, Coprococcus, Dialister, Eubacterium, Lachnospiraceae, Olibacterium, Roseburia, Christensenellaceae, Erwinia, Flavonifracter, Oscillospira, Phascolactobacterium, Prevotella, Succinivibrio, Ruminococcus and / or Veillonella.

[0017] Additionally, lipogenic bacterial species include, but are not limited to, Acinetobacter baumannii, Bacillus cereus, Bacillus anthracis, Bacillus subtilis, Bacteriodes thetaiotaomicron, Valteriodes vulgatus, Bartonella henselae, Bordetella pertussis, Borrelia recurrentis, Borrelia ermsii, Borrelia tulicachue, Borrelia burgdorferi, Campylobacter jejuni, Citrobacter fluendi, Chlamydia psittaci, Chlamydia trachomatis, Chlamydia pneumoniae, Clostridium botulinum, Clostridium difficle, Clostridium tetani, Clostridium perfringens, Clostridium ramosum, Clostridium novyi, Clostridium septicum, Clostridium leptum, Corynebacterium dyslipidae, Desulfovibrio piger, Ehrlichia chaffeensis, Enterococcus faecalis, Escherichia coli (e.g. EHEC, EIEC, ETEC), Faecalibacterium prausnitzii, Francisella tularensis, Helicobacter pylori, Haemophilus influenzae, Haemophilus parainfluenzae, Haemophilus aegyptus, Klebsiella pneumoniae, Lactobacillus reuteri, Legionella pneumophila, Leptospirex erythroderma, Leptospira icterolae, Listeria monocytogenes, Methanobrevibacter smithii, Moraxella catarrhalis, Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacter Mycobacterium asiaticum, Mycobacterium intracellulare, Mycobacterium avium intracellulare, Myobacterium johnnei, Mycobacterium avium, Mycobacterium smegmatis, Mycoplasma pneumoniae, Mycoplasma hominis, Neisseria meningitidis, Neisseria gonorrhoeae, Rickettsia prowozekii, Rickettsia rickettsii, Rickettsia akari, Propionibacterium acnes, Pseudomonas aeruginosa, Pseudomonas syringae, Salmonella typhi, Salmonella typhi, Salmonella paratyphi, Salmonella schott-mulleryi, Salmonella hirskelii, Shigella dysenteriae, Spirirum minus, Staphylococcus aureus, Staphylococcus epidermidis, Stenotrophomonas maltophilia,Streptobacillus moniliformis, Streptococcus pneumoniae, Streptococcus mutans, Streptococcus oralis, Streptococcus parasanguis, Streptococcus pyogenes, Streptococcus viridans, Streptococcus coelicolor, Streptococcus agalactiae, Streptococcus bovis, Treponema pallidum, Treponema pertainae, Treponema calateum, Vibrio cholerae, Vibrio parahaemolyticus, Yersinia pestis, E. The bacterial species is selected from Luscinia enterocolitica, Bacteroides fragilis, Blautia hydrogenotrophica, Coprococcus catus, Eubacterium ventriosum, Bacteroides faecitinetillae, Bifidobacterium animalis, Blautia wechslerae, Clostridium volteae, Flavonifracter proutii, Lactobacillus gasseri, Ruminococcus brommii and / or Ruminococcus obeum, where each bacterial species is presumed to have a known and / or suspected adipogenic effect on humans and / or animals.

[0018] In the first step of the method according to the invention, a biological sample containing bacteria of at least one adipogenic bacterial species must be provided. The term "biological sample" relates to human and / or animal material and / or material exposed to humans and / or animals for at least a short period of time. In this context, it has been found essential that the biological sample contains bacteria of at least one adipogenic bacterial species, preferably 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more bacterial species. Samples of this kind are well known to the skilled person, such as cell or nucleus-containing (DNA and / or RNA-containing) samples of saliva, urine, blood, feces, sweat, tissue, organ puncture, organs, parts of organs, whole organisms (including dead / necrotic), soil samples, water samples or similar samples, among others.

[0019] The next step of the method according to the invention requires providing bacteriophages of at least one bacteriophage species that are specific for at least one lipogenic bacterial species. The term "bacteriophage" or "bacteriophage species" is understood by those skilled in the art and relates to an infectious organic structure that can spread outside the cell and grow inside the cell. The cell acts as a host and, in the case of bacteriophages, they are essentially bacterial and have a pronounced host specificity, i.e., they are specific for at least one bacterial species and use only the bacteria that they are specific for spreading and growing.

[0020] It is important that the bacteriophage used in the methods of the present invention comprises at least one nucleic acid functionally linked to a promoter and / or regulatory element or has a promoter and / or regulatory element associated with a nucleic acid, said nucleic acid encoding either an antimicrobial nucleic acid molecule, a bacterial polypeptide and / or a fragment thereof. Preferably, the bacteriophage comprises a nucleic acid having at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more promoters and / or regulatory elements. Nucleic acid sequences encoding promoters or regulatory sequences are well known to those skilled in the art.

[0021] Those of skill in the art are well aware that the terms "nucleic acid," "gene," "DNA," "RNA," "mRNA," "cRNA," "miRNA," and the compounds "nucleic acid sequence" and "nucleic acid molecule" are used interchangeably and synonymously to describe either single- or double-stranded deoxyribonucleotides and ribonucleotides and polymers thereof.

[0022] The terms "promoter" and "regulatory sequence" and / or "regulatory element" are also familiar to the person skilled in the art and relate to a nucleic acid sequence of DNA that allows and / or increases the expression, i.e. the transcription of a gene into mRNA. In this case, enhancing elements, i.e. binding sites for growth factors, hormones, oncogenes and / or the like, may be regulatory elements.

[0023] Those skilled in the art are further familiar with the terms "polypeptide", "peptide", "amino acid" and "protein", which are used interchangeably and synonymously to refer to a polymer of amino acid residues. Naturally occurring amino acids are those encoded by the genetic code as well as those amino acids that may be later modified. Polypeptides may exist as residual substances, as transport and / or signaling molecules, as structural molecules, as protective and / or defense molecules, and / or as metabolically active molecules, and may perform different functions inside and / or outside a cell and / or organism. Polypeptides may, for example, have antibacterial effects, interfering with bacterial ecology due to their properties or functions.

[0024] Within the scope of the method according to the invention, at least one antimicrobial nucleic acid molecule, at least one antimicrobial polypeptide, and / or at least one fragment thereof is provided; preferably, the bacteriophage may code for 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more antimicrobial nucleic acid molecules, antimicrobial polypeptides and / or fragments thereof. The antimicrobial nucleic acid molecule may be derived from a region of an RNA species, for example a functional mRNA or miRNA. The polypeptide may exist as a simple linear polypeptide or may also be a folded structural protein. Other acceptable proteins may also be molecules that act as toxins, growth inhibitors and enzymes or molecules for inhibiting cell division. Acceptable nucleic acid molecules, for example RNA molecules, are those that inhibit metabolic processes, inhibit cell division or promote the malfunction of biological processes in the bacterium.

[0025] Within the scope of the methods according to the present invention, nucleic acid sequences encoding nucleic acid molecules, polypeptides and / or fragments thereof are considered, which are at least 50% identical to the above-mentioned antimicrobial nucleic acid sequences or antimicrobial polypeptides. Preferably, the nucleic acid sequences are also considered to be at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identical to the above-mentioned antimicrobial nucleic acid sequences or antimicrobial polypeptides. In this context, the nucleic acid sequences are, for example, naturally occurring or non-naturally occurring nucleic acid sequences already present in the bacteriophage or introduced into the respective bacteriophage via molecular biology methods. For example, these molecular biology methods may refer to in vivo and / or in vitro recombination, gene transfer, CRISPR / Cas, TALEN, etc.

[0026] Said bacteriophage may belong to the classes Caudovirales, Ligamenviralis etc. Preferably, the bacteriophage belongs to the family Ackermannviridae, Ampulaviridae, Bicaudaviridae, Clavaviridae, Corticoviridae, Fuselloviridae, Globuloviridae, Guttaviridae, Lipotryxviridae, Myoviridae, Plasmaviridae, Podoviridae, Portogloboviridae, Ruziviridae, Saltaproviridae, Spherolipoviridae, Siphoviridae, Techtiviridae, Tristomaviridae, Turiviridae, Inoviridae, Microviridae, Spiraviridae, Pleolipoviridae, Cystoviridae or Leviviridae. It is contemplated within the scope of the present invention that bacteriophages of at least one, preferably 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more bacteriophage species are provided. Furthermore, it is contemplated that the bacteriophages of a bacteriophage species are specific for at least one, preferably 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more lipogenic bacterial species. Target selection of suitable bacteriophages for the corresponding bacterial species is well known to the skilled person and can be easily selected, for example, from a data bank.

[0027] The next step of the method according to the invention involves exposing the biological sample to the bacteriophage and incubating them together. The biological sample and the bacteriophage can be exposed to each other by dropping, injecting, placing on top of each other, and / or through different methods known to those skilled in the art.

[0028] The term "incubation" in the context of the method according to the invention relates to a method step in which a biological sample is incubated with a bacteriophage for a certain period of time; preferably under corresponding circumstances well known to the skilled person, the result is a regeneration cycle of the bacteriophage in the biological sample containing bacteria of at least one bacterial species specific for the bacteriophage. In addition to incubating the biological sample with the bacteriophage, this method step also includes a step of monitoring the biological sample with the bacteriophage until the population of at least one bacterial species is reduced by at least 70%. The implementation of the monitoring is well known to the skilled person and can be carried out, for example, by observing the sample for a predetermined period of time or based on the experience of the skilled person. Alternatively, a colored or other visually perceptible indicator can be intended that signals when the population of the bacterial species is reduced by at least 70%. This can occur, for example, by chemical and / or biochemical indicators, such as when a desired bacterial density is achieved or by coloring, a color change or color loss of the coloring. The incubation and monitoring steps can be performed one after the other until the bacterial species is reduced by at least 70%. This may occur, for example, when a desired bacterial density is achieved, or by chemical and / or biochemical indicators such as staining, a change in staining color, or loss of color, etc. The incubating and monitoring steps may be performed one after the other until the bacterial species is reduced by at least 70%.

[0029] Within the scope of the present invention, it has been proven to be advantageous if the population of bacterial species is reduced by at least 70%.Preferably, the population can be reduced by more than 70%, for example at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100%; a reduction of at least 70% of the population of bacterial species is considered substantial, and a reduction of 95% of the population of bacterial species is considered significant.Those skilled in the art are well aware that a reduction of 100% is rare, even though the use of suitable bacteriophage in suitable biological sample can bring about this result.

[0030] With a 70% to 100% reduction in the population of bacterial species, the skilled artisan may further recognize the success of the method to account for sufficient antibacterial effect and sufficient reduction of adipogenic bacterial species in a biological sample. The reduction in the population of bacterial species may be used as a metric for the effectiveness or success of a weight loss treatment based on the use of the respective bacteriophage. Alternatively, the reduction in bacterial species may be targeted to counteract the life or ecology of the anti-adipogenic bacterial species. As a result, the anti-adipogenic bacterial species may continue to persist or grow while the bacterial species that counteract the anti-adipogenic bacterial species is reduced, even though the anti-adipogenic bacterial species is specific to the bacteriophage. In this context, it is speculated that the result of the growth of the population of anti-adipogenic bacterial species may have the same effect on the organism as the reduction in the population of adipogenic bacterial species.

[0031] The method according to the invention allows the population of at least one adipogenic bacterial species to be specifically, effectively, reciprocally, medically safe, permanently and persistently reduced in a biological sample, without damaging and / or affecting bacteria or other cells, tissues and / or organs in the same biological sample for which the bacteriophage has no specificity, and without any side effects. In this way, in contrast to the use of antibiotics, damage to other bacterial species, such as yeasts, tissues and / or organs in humans and / or animals, is completely avoided. The method according to the invention can be performed quickly and easily, as well as reliably and cost-effectively, since a small number of method steps can be performed in a standard manner, and a high success rate is achieved by a considerable (70% or more) or significant (95% or more) reduction of the bacterial species. The method can furthermore be performed using simple means, requiring only equipment familiar to the skilled person for providing the biological sample and the bacteriophage, exposing and incubating them, and for evaluating the reduction of the respective bacterial species in the population.

[0032] Advantageous further embodiments of the invention, which can be realized individually or in combination, are set out in the dependent claims.

[0033] According to a further embodiment of the invention, step c) is followed by a step in which the reduction of the population of lipogenic bacterial species is evaluated. The term "evaluate" is clear to the skilled person and relates to the analysis, evaluation and / or diagnosis of step c). Since a reduction of the population of bacterial species of at least 70% is monitored, this method step serves in particular to verify the reduction achieved, as well as to record the results by preparing written notes, photographs and video recordings, to obtain and document data on enumeration, density measurement, growth determination or similar methods, which can be used to generate statistics on repeated implementations of the method according to the invention. In this step, the vitality and / or viability of bacteria remaining in the biological sample and not killed or not sufficiently killed by the bacteriophage can be established. In this context, an evaluation of the vitality of the bacteria for which the respective bacteriophage is specific is possible. General methods for this purpose are also well known to the skilled person.

[0034] It is further conceivable that the nucleic acid has more than one promoter and / or regulatory element in step b), preferably there are 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more promoter and / or regulatory elements. In this context, it is important to achieve a particularly strong activation of the expression of the nucleic acid and an increased and efficient production of the antimicrobial nucleic acid, antimicrobial nucleic acid molecule, antimicrobial polypeptide and / or fragments thereof. Bacteriophages carrying such nucleic acids may contribute particularly efficiently to the reduction of the population of bacterial species, for example considerably accelerating and / or considerably shortening step c) (incubation) of the method according to the invention.

[0035] In yet another further embodiment, it is envisaged that the nucleic acids and nucleic acid sequences code for two or more nucleic acid molecules, polypeptides and / or fragments thereof, the nucleic acids preferably coding for 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more nucleic acid molecules, polypeptides and / or fragments thereof. These nucleic acid molecules, polypeptides and / or fragments thereof may have an antibacterial effect on one or more bacterial species; it is preferred that a bacteriophage carrying such a nucleic acid may infect several bacterial species. Furthermore, it may be intended that the nucleic acid sequence is specific for killing one bacterial species. For this purpose, the method may be accelerated and a substantial or significant reduction in the population of bacterial species may be achieved within a biological sample. This increases the efficiency of the method while also reducing the duration and / or cost of carrying out the method.

[0036] In another embodiment of the method, it is contemplated that the adipogenic bacterial species is anaerobic and / or aerobic, and that the bacterial species originates from a strain of Actinobacteria, Bacteroidetes, Firmicutes or Proteobacteria.

[0037] The terms "aerobic" and "anaerobic" are well known to those skilled in the art and describe the preferred lifestyle of the respective organisms. In this context, an "aerobic" lifestyle is understood to mean that the organism depends on oxygen in the air, or rather on oxygen in the air for survival. In contrast, "anaerobic" refers to a lifestyle that survives without oxygen, meaning that the respective organism can maintain its molecular metabolism without oxygen. A distinction is made between "obligately", "permissively" and "facultatively" aerobic and / or anaerobic organisms, and the skilled artisan will understand that "obligate" exclusively describes an aerobic or anaerobic lifestyle, to the exclusion of the respective other lifestyle. However, "permissive" means that the organism lives aerobically or anaerobically, but can tolerate the opposite oxygen conditions, at least for a certain period of time. Finally, the term "facultative" means that an anaerobic organism, for example, generally maintains the corresponding metabolic processes, but can switch to aerobic metabolic processes when presented with oxygen, and vice versa. Thus, bacterial species may have an exclusively aerobic or anaerobic lifestyle, or a combination thereof.

[0038] The above term definitions and / or embodiments are presumed to apply to all aspects described in the following description unless otherwise indicated.

[0039] According to the present invention there is further proposed a kit for carrying out the method according to the invention, said kit comprising: a) a container containing a bacteriophage of at least one bacteriophage species, the bacteriophage being specific for at least one adipogenic bacterial species and comprising at least one nucleic acid operably linked to a promoter and / or a regulatory element; and b) instructions for carrying out said method; Equipped with.

[0040] The term "kit" as used herein refers to a kit-of-parts of the aforementioned components, which are preferably provided separately or in individual containers.

[0041] The term "instructions for carrying out said method" relates to numerical, descriptive and / or graphical representations of instructions that allow carrying out the objectives of said method in a simplified manner. The instructions are preferably available in the form of a handbook or may be provided via a computer program such as an application. Furthermore, it is conceivable that the computer program has implemented algorithms that are capable of carrying out the discriminations, comparisons and / or their results referred to in the method of the invention at hand. The computer program may be made available on a device such as a data storage medium or an optical storage medium (e.g. a compact disc) or directly on a computer or data processing device. In addition, said instructions may preferably comprise criteria regarding the amounts used, as known to the person skilled in the art. Furthermore, said instructions include the necessary conditions for the storage and disposal of the container and the biological sample. Furthermore, the instructions for carrying out the method may include tables, registers, databanks and excerpts of the same regarding adipogenic, antiadipogenic and / or other bacterial species and bacteriophage species known to the person skilled in the art; regarding bacteriophage species, information regarding their specificity and / or efficiency may also be listed.

[0042] According to the present invention, there is further proposed a bacteriophage comprising at least one nucleic acid functionally linked to a promoter and / or a regulatory element, said bacteriophage being specific for at least one lipogenic bacterial species, said nucleic acid being selected from the group: a) a nucleic acid sequence encoding at least one antimicrobial nucleic acid molecule; and b) a nucleic acid sequence encoding a nucleic acid molecule, said nucleic acid sequence being at least 50% identical to the nucleic acid molecule encoded by the nucleic acid sequence in a); and c) a nucleic acid sequence encoding at least one antimicrobial polypeptide; and d) a nucleic acid sequence encoding a polypeptide, said nucleic acid sequence being at least 50% identical to the polypeptide encoded by the nucleic acid sequence in c); and e) a nucleic acid sequence for a fragment of a nucleic acid from a), b), c), or d), encoding a nucleic acid molecule or polypeptide. is selected from among:

[0043] The bacteriophage according to the invention has been described in detail above, and said nucleic acid sequence is a naturally occurring nucleic acid sequence or a non-naturally occurring nucleic acid, which is already present in the bacteriophage or which has been introduced into the bacteriophage via molecular biology methods. For example, these molecular biology methods may involve in vivo and / or in vitro recombination, gene transfer, CRISPR / Cas, TALEN, etc., as known to the skilled person.

[0044] In a further embodiment, it is conceivable that the nucleic acid has two or more promoters and / or regulatory elements. It is important that a particularly strong activation of the expression of the nucleic acid and a particularly increased and efficient production of the antibacterial nucleic acid, antibacterial nucleic acid molecule, antibacterial polypeptide and / or fragments thereof can be achieved. Bacteriophages carrying such nucleic acids can contribute particularly efficiently to the reduction of the population of lipogenic bacterial species or to the reduction of bacterial species that counteract the ecology of antilipogenic bacterial species.

[0045] It is further contemplated that said nucleic acid may encode nucleic acid sequences for more than one nucleic acid molecule, polypeptide and / or fragment thereof, in which case the bacteriophage is preferably able to use such nucleic acid to infect several bacterial species.

[0046] In yet another embodiment, the adipogenic bacterial species is anaerobic and / or aerobic, and the bacterial species may originate from a strain of Actinobacteria, Bacteroidetes, Firmicutes or Proteobacteria.

[0047] Further according to the present invention bacteriophages and / or pharmaceutical compositions comprising a bacteriophage and at least one other component are proposed for use in medicaments for altering the intestinal flora, for achieving and / or maintaining weight loss, for preventing weight gain and / or for treating overweight, obesity, metabolic disorders, cardiovascular diseases, bacterial infections, metabolic diseases, metabolic syndrome and / or cancer.

[0048] The term "pharmaceutical composition" as used herein refers to a mixture of a bacteriophage as described in more detail above with at least one other ingredient. Preferably, such other ingredients may be stabilizers, wetting agents, pharmaceutical carriers, pharma- ceutically acceptable carriers, diluents, pharma- ceutical acceptable diluents, further pharma- ceutical active ingredients, release agents, and the like. Preferred diluents are water, alcohol, saline, buffers such as phosphate buffered saline, syrups, oils, water, emulsions, different types of wetting agents, and the like. The carrier must be acceptable in the sense of being compatible with the other ingredients of the composition and not harmful to humans and / or animals. The pharmaceutical carrier used may contain a solid, gel, or liquid. Examples of solid carriers include lactose, terra alba, sucrose, talcum powder, gelatin, agar, pectin, acacia, magnesium stearate, stearic acid, and the like. In a similar manner, the carrier or diluent may contain a time-delay material well known in the art, such as glycerol monostearate or glycerol distearate, by itself or with a wax. These suitable carriers include the above-mentioned carriers and other carriers known in the art, such as those mentioned in Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pennsylvania; the European Pharmacopoeia; the Homeopathic Pharmacopoeia of the United States; or the Homeopathic Pharmacopoeia (HAB). Pharmaceutically acceptable diluents are selected so that the biological activity of the combination is not affected. Examples of such diluents include distilled water, saline, Ringer's solution, dextrose solution and HANK solution. Pharmaceutical compositions may also contain other carriers, additives or non-toxic, non-therapeutic, non-immunogenic stabilizers, etc.

[0049] The bacteriophage and / or pharmaceutical composition should be adapted to each specific use. It can therefore be assumed that the bacteriophage and / or pharmaceutical composition may be formulated for systemic or local application depending on the intended administration. Preferably, the bacteriophage and / or pharmaceutical composition should be formulated for systemic or local administration. Preferably, oral administration, for example in the form of a tablet, solution or drinkable ampoule, or topical administration in gel form, or administration by injection, is intended. However, depending on the type and mode of action, the bacteriophage and / or pharmaceutical composition may be administered in different ways, including cutaneous, intramuscular, subcutaneous, oral, rectal, retrograde or intravenous administration. In principle, the exact, individually proposed dosage may depend on further parameters well known to experts in the field. For example, children may receive a different dose than adults. Using the various calculation tools at their disposal, medical professionals can easily determine whether the dosage needs to be adapted.

[0050] The term "change in the composition of gut microbiota" describes a process that can be understood by a person skilled in the art, in which at least one population of lipogenic bacterial species that is a component of the gut microbiota of humans and / or animals is increased, decreased and / or affected in other ways. This can be measured by isolating and reviewing to determine the number of corresponding bacteria, or it can be determined by comparing with other bacterial species in the gut microbiota. Appropriate methods are common knowledge to a person skilled in the art.

[0051] The term "achieving and / or maintaining weight loss" is understandable to the skilled artisan and relates to a short-term reduction in the body weight of a human and / or animal and / or preferably a sustained maintenance of the weight loss in a human and / or animal. The term "preventing weight gain" is also understandable to the skilled artisan and relates to a sustained and / or continuous maintenance of the weight loss in a human and / or animal. In this case, achieving and / or maintaining weight loss and prevention of weight gain in a human and / or animal is preferably achieved by reducing at least one adipogenic bacterial species.

[0052] The use of bacteriophages and / or pharmaceutical compositions in medicine may occur, inter alia, for treating diseases in oncology, immunology, infectology, oral and maxillofacial surgery, otorhinolaryngology, ophthalmology, neurology, gynecology, gastroenterology, endocrinology, psychiatry, psychosomatics, orthopedics, pediatrics, surgery, urology, and / or similar medical fields. Acceptable diagnoses for which the use of the bacteriophage and / or pharmaceutical composition may be indicated may be, inter alia, metabolic syndrome, type 2 diabetes, gallbladder diseases, chronic diseases of the digestive tract, chronic intestinal inflammation, hypochlorhydria, hypertension, lipid metabolism disorders, respiratory depression, sleep apnea, coronary heart disease, arthropathy, gout, cancer such as uterine, breast, cervical, pancreatic, liver, stomach, intestine, prostate and gallbladder cancer, sex hormone disorders, decreased libido, joint and back pain, increased risk of thrombosis and embolism, increased risks during surgery and narcotics, psychosocial problems and resulting limitations in quality of life resulting from depression caused by reduced self-esteem and / or the perception of being less valued by the environment, etc. Preferably, the aim in each case is to reduce at least one lipogenic bacterial species, leading to the achievement and / or maintenance of, as a primary or secondary success, weight loss and / or prevention of weight gain in humans and / or animals, in order to obtain a good, preferably improved, prognosis for the treatment of the respective disease and / or the respective affliction. What is considered is the use of the bacteriophage and / or pharmaceutical composition independent of the actual disease. The treatment is considered successful if at least one lipogenic bacterial species or weight is reduced and / or if weight gain is stopped or delayed.

[0053] The term "treatment" relates to the respective improvement of the human and / or animal that occurs compared to an untreated human and / or animal, this improvement being preferably based on a reduction in at least one lipogenic bacterial species, achieving and / or maintaining and / or preventing weight loss in the human and / or animal. It is presumed that in 100% of the treated humans and / or animals, the treatment is probably not successful. However, the term presupposes that the treatment was successful for a statistically significant portion of the subjects (e.g. a cohort in a cohort study). The skilled person can easily establish whether a portion is statistically significant using different known statistical evaluation means, for example, the determination of confidence intervals, the determination of p-values, Student's t-test, Mann-Whitney U-test, etc. Preferred confidence intervals are at least 90%, at least 95%, at least 97%, at least 98% or at least 99%. The p-value is preferably 0.05; 0.01; 0.005 or 0.0001.

[0054] The term "medicament" refers to a therapeutically effective amount of the bacteriophage and / or pharmaceutical composition as described above. Preferably, the pharmaceutical composition comprises the bacteriophage, at least one pharma- ceutically acceptable carrier and / or diluent. The medicament may be formulated for different modes of administration as detailed above. The therapeutically effective amount refers to the amount necessary to change the composition of the intestinal microflora, to reduce at least one lipogenic bacterial species, to achieve and / or maintain weight loss, to prevent weight gain, and / or to treat the above-mentioned diseases. The therapeutic efficacy and toxicity may be determined via pharmaceutical standard methods in cell cultures or test animals, such as ED50 (the dose that is therapeutically effective in 50% of cases) and LD50 (the dose that is lethal in 50% of cases). The dose ratio between therapeutic and toxic effects is the therapeutic index, which can be expressed as the LD50 / ED50 ratio. The dosing regimen is determined by the physician and other clinical factors. As is known in the medical field, the corresponding dosage depends on many factors known to medical professionals, including height, body surface area, age, substance administered, sex, time and mode of administration (general health condition and other drugs administered at the same time). Progress can be monitored by regular evaluation.

[0055] Furthermore, the bacteriophage according to the present invention and / or a pharmaceutical composition comprising a bacteriophage and at least one other component are proposed for use in the treatment of overweight, obesity, metabolic disorders, cardiovascular disease, bacterial infections, metabolic diseases, metabolic syndrome and / or cancer.

[0056] Further according to the present invention, bacteriophages and / or pharmaceutical compositions comprising a bacteriophage and at least one other component are proposed for use in therapeutic and / or non-therapeutic methods for altering the composition of the intestinal microbiota, for achieving and / or maintaining weight loss, for preventing weight gain and / or for treating overweight, obesity, metabolic disorders, cardiovascular diseases, bacterial infections, metabolic diseases, metabolic syndrome and / or cancer.

[0057] In another embodiment of the invention, the bacterial infection is considered to be a bacterial infection of the respiratory tract, teeth, mouth, jaw, eyes, musculoskeletal system, blood, gastrointestinal tract, preferably the large intestine, small intestine, duodenum, stomach, liver, gallbladder and / or pancreas, skin, cardiovascular system, hormonal balance, psychiatry, immune system, nervous system, metabolic system, wounds and / or genitourinary tract.

[0058] The term "gastrointestinal tract" is well known to those skilled in the art and relates to the organ in humans and / or animals that functions in the intake, breakdown, transport and / or processing of nutrients in order to make them available via the human and / or animal organ, the intestine, in particular the intestinal flora, and contributes significantly to their regulation.

[0059] The described features can be realized either alone or in combination with each other, however, the present invention is not limited to the exemplary embodiments contained herein.

Claims

1. 1. A method for reducing the population of at least one adipogenic bacterial species, comprising the steps of: a) providing a biological sample containing bacteria of at least one adipogenic bacterial species; and b) providing a bacteriophage of at least one bacteriophage species specific for at least one adipogenic bacterial species and comprising at least one nucleic acid operably linked to a promoter and / or a regulatory element, said nucleic acid being selected from the group consisting of: i) a nucleic acid sequence encoding at least one antimicrobial nucleic acid molecule; and ii) a nucleic acid sequence encoding a nucleic acid molecule, said nucleic acid sequence being at least 50% identical to the nucleic acid molecule encoded by the nucleic acid sequence in i; and iii) a nucleic acid sequence encoding at least one antimicrobial polypeptide; and iv) a nucleic acid sequence encoding a polypeptide, said nucleic acid sequence being at least 50% identical to the polypeptide encoded by the nucleic acid sequence in iii; and v) nucleic acid sequences for fragments of nucleic acids, nucleic acid molecules or polypeptide-encoding fragments from i, ii, iii, iv. the step selected from: c) exposing the biological sample to the bacteriophage and incubating it with the bacteriophage, said incubation being carried out until the population of the lipogenic bacterial species is reduced by at least 70%. The method comprising:

2. The method of claim 1 , wherein following step c) the reduction in the population of adipogenic bacterial species is assessed.

3. The method of claim 1 or 2, wherein the nucleic acid has two or more promoters and / or regulatory elements.

4. The method according to any one of claims 1 to 3, wherein the nucleic acid encodes the nucleic acid sequences of two or more identical nucleic acid molecules, polypeptides and / or fragments.

5. The method according to any one of claims 1 to 4, wherein the adipogenic bacterial species is anaerobic and / or aerobic, said bacterial species may be attributed to a strain of Actinobacteria, Bacteroidetes, Firmicutes or Proteobacteria.

6. a) a container containing a bacteriophage of at least one bacteriophage species, the bacteriophage being specific for at least one adipogenic bacterial species and comprising at least one nucleic acid operably linked to a promoter and / or a regulatory element; and b) instructions for carrying out said method: A kit for carrying out the method according to any one of claims 1 to 5, comprising:

7. A bacteriophage comprising at least one nucleic acid operably linked to a promoter and / or a regulatory element, said nucleic acid being specific for at least one group of adipogenic bacteria, said nucleic acid being specific for at least one group of adipogenic bacteria: a) a nucleic acid sequence encoding at least one antimicrobial nucleic acid molecule; and b) a nucleic acid sequence encoding a nucleic acid molecule, said nucleic acid sequence being at least 50% identical to the nucleic acid molecule encoded by the nucleic acid sequence in a); and c) a nucleic acid sequence encoding at least one antimicrobial polypeptide; and d) a nucleic acid sequence encoding a polypeptide, said nucleic acid sequence being at least 50% identical to the polypeptide encoded by the nucleic acid sequence in c); and e) a nucleic acid sequence for a nucleic acid fragment, nucleic acid molecule or polypeptide-encoding fragment from a), b), c), d), or e). The bacteriophage is selected from the group consisting of

8. The bacteriophage of claim 7 , wherein the nucleic acid has two or more promoters and / or regulatory elements.

9. 9. A bacteriophage according to claim 7 or 8, wherein the nucleic acid encodes the nucleic acid sequences of two or more identical nucleic acid molecules, polypeptides and / or fragments.

10. A bacteriophage according to any one of claims 7 to 9, wherein the adipogenic bacterial species is anaerobic and / or aerobic, and the bacterial species may be attributed to a strain of Actinobacteria, Bacteroidetes, Firmicutes or Proteobacteria.

11. 11. A bacteriophage according to any one of claims 7 to 10 and / or a pharmaceutical composition comprising a bacteriophage according to any one of claims 7 to 10 and at least one other ingredient for use in a medicament for altering the gut flora, for achieving and / or maintaining weight loss, for preventing weight gain and / or for treating overweight, obesity, metabolic disorders, cardiovascular disease, bacterial infections, metabolic discomfort, metabolic syndrome and / or cancer.

12. 11. A bacteriophage according to any one of claims 7 to 10 and / or a pharmaceutical composition comprising a bacteriophage according to any one of claims 7 to 10 and at least one other ingredient for use in a medicament for treating overweight, obesity, metabolic disorders, cardiovascular disease, bacterial infections, metabolic discomfort, metabolic syndrome and / or cancer.

13. 11. A bacteriophage according to any one of claims 7 to 10 and / or a pharmaceutical composition comprising a bacteriophage according to any one of claims 7 to 10 and at least one other ingredient for use in a therapeutic or non-therapeutic method for altering the composition of the gut microbiota, for achieving and / or maintaining weight loss, for preventing weight gain and / or for treating overweight, obesity, metabolic disorders, cardiovascular diseases, bacterial infections, metabolic discomfort, metabolic syndrome and / or cancer.

14. 14. The bacteriophage and / or pharmaceutical composition according to any one of claims 11 to 13, wherein the bacterial infection is a bacterial infection of the respiratory tract, teeth, mouth, jaw, eyes, musculoskeletal system, blood, gastrointestinal tract, skin, cardiovascular system, hormone balance, psychiatry, immune system, nervous system, metabolism, wounds and / or urogenital tract.