Compositions and methods for characterizing the microbiome

Abstract

A system is provided that includes a plurality of C. elegans cultures, each culture containing a transgenic C. elegans strain that models a mammalian disease or pathology. Methods of using the system include, for example, characterizing microbial strains of the mammalian microbiome and determining whether such microbial strains affect a mammalian disease or disorder.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Patent Application No. 62 / 899,718, filed September 12, 2019, and U.S. Provisional Patent Application No. 62 / 988,132, filed March 11, 2020, the entire contents of which are incorporated herein by reference in their entirety. [Background technology]

[0002] Caenorhabditis elegans is a bacteriovorus nematode that is approximately 1 mm in length and lives in temperate soil environments. Summary of the Invention

[0003] The present disclosure provides insight that C. elegans may provide and / or represent a surprisingly useful system for assessing one or more characteristics of a microbial preparation (e.g., of a microbiome sample). In particular, the present disclosure describes techniques that may be useful for assessing a microbiome sample to identify or characterize the influence and / or regulation of the microbial strains of such a microbiome sample on a particular disease or condition. In some embodiments, such techniques may be useful for identifying strain-level differences in a particular patient or patient population. Thus, the present disclosure also provides techniques that may be useful for assessing the nature of the microbial strains in a patient-specific sample, and thus for providing patient-specific information on how an individual patient's microbiome differentially impacts the patient's health status. For example, in some embodiments, the techniques provided herein may be useful for identifying a disease or condition to which a patient may be susceptible based on the nature of the microbial strains in a patient-specific sample. In some embodiments, the techniques provided herein may be useful for identifying microbial strains in a patient that are beneficial, for example, for protecting the patient from or conferring resistance to a particular disease or condition. Thus, the technology described herein is useful as a diagnostic tool for screening microbiome samples (e.g., human microbiome samples) for disease modifying agents (e.g., microbial strains that affect a disease or pathology).

[0004] Indeed, in certain embodiments, the use of a transgenic C. elegans whole animal model system to identify or screen for microbiome strains (e.g., present in the human microbiome) that may modulate or affect the pathogenesis and / or development of a neurodegenerative disease or condition (e.g., Alzheimer's disease). Upon review of this disclosure, one of skill in the art will appreciate that the technology described herein is applicable for use in connection with neurodegenerative diseases such as Alzheimer's disease, as exemplified above, as well as in connection with a variety of other diseases or conditions that may be associated with the microbiome, such as, but not limited to, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis, type II diabetes, obesity, hyperglycemia, impaired glucose tolerance, insulin resistance (i.e., hyperinsulinemia, metabolic syndrome, syndrome X), hypercholesterolemia, hypertension, hyperlipoproteinemia, hyperlipidemia (e.g., dyslipidemia), hypertriglyceridemia, cardiovascular disease, atherosclerosis, peripheral vascular disease, kidney disease, ketoacidosis, thrombotic disorders, nephropathy, diabetic neuropathy, diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, glaucoma, sexual dysfunction, dermatopathy, dyspepsia, hypoglycemia, metabolic syndrome, cancer, or edema.

[0005] In some aspects, provided herein is a system comprising a plurality of C. elegans cultures, each culture comprising a transgenic C. elegans strain that models a disease or condition (e.g., a mammalian disease or condition). In some embodiments, such a plurality of C. elegans cultures may comprise at least 5 or more (e.g., including at least 6, at least 7, at least 8, at least 9, at least 10, at least 15, at least 20, at least 25, at least 30, at least 40, at least 50, or more) C. elegans cultures.

[0006] In some embodiments, one or more of the C.elegans cultures in the provided system comprises a transgenic C.elegans strain that models a disease or condition present in the target subject. In some embodiments, one or more of the C.elegans cultures in the provided system may comprise a transgenic C.elegans strain that models a mammalian disease or condition (e.g., a human disease or condition). Exemplary human diseases or conditions that the C.elegans strain models may include neurodegenerative diseases or disorders (e.g., Alzheimer's disease). In some embodiments, the diseases or conditions that the C.elegans strain models (e.g., a human disease or condition) may include diseases or conditions associated with an altered or defective HIF-1 pathway (e.g., may include a cellular stress response). In some embodiments, for example, the disease or condition that the C.elegans models may be an intraocular neovascular disease or disorder (e.g., diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma). In some embodiments, one or more of the C. elegans cultures in the provided systems may include transgenic C. elegans that model a non-human mammalian disease or condition, such as a canine, feline, equine, bovine, ovine, caprine, or porcine disease or condition.

[0007] In some embodiments, the transgenic C. elegans strains provided in the systems described herein may include a transgene that includes a characteristic sequence element associated with a target disease or condition (e.g., a mammalian disease or condition). Such characteristic sequence elements associated with a disease or condition (e.g., a mammalian disease or condition) may be or include a foreign gene (e.g., a mammalian gene), a DNA regulatory element (e.g., a mammalian DNA regulatory element), and / or a mammalian RNA regulatory element. A variety of DNA and RNA regulatory elements are known in the art, and thus, one of ordinary skill in the art will understand that in some embodiments, a DNA regulatory element associated with a disease or condition (e.g., a mammalian disease or condition) may be or include an enhancer, promoter, silencer, insulator, locus control region, and combinations thereof. In some embodiments, RNA regulatory elements associated with a disease or condition (e.g., a mammalian disease or condition), such as, for example, untranslated regions, introns, splice sites, and combinations thereof, may be used in accordance with the present disclosure.

[0008] Additionally or alternatively, the transgenic C. elegans strains in the provided systems may contain transgenes, including reporter genes, non-limiting examples of which may include, but are not limited to, fluorescent, phosphorescent, and / or bioluminescent proteins.

[0009] In some embodiments involving C. elegans cultures, at least two or more of such cultures may each comprise a transgenic C. elegans strain that models the same disease or condition (e.g., a mammalian disease or condition). In some such embodiments, at least two or more of such cultures may each comprise a transgenic C. elegans strain that models a different biochemical or molecular pathway associated with the same disease or condition (e.g., a mammalian disease or condition). In some embodiments involving C. elegans cultures, all of such cultures may each comprise the same transgenic C. elegans strain that models the same disease or condition (e.g., a mammalian disease or condition).

[0010] In some embodiments involving C. elegans cultures, at least two or more of such cultures may each comprise a transgenic C. elegans strain that models a different disease or condition (eg, a mammalian disease or condition).

[0011] The systems described herein can be used to characterize the relationship and / or effect of a microbial preparation (e.g., of a microbiome sample) with a certain disease or condition in a target subject. Thus, in some embodiments, one or more C. elegans cultures in such systems comprise microbes of a mammalian microbiome (e.g., a human microbiome). In some embodiments, each of such C. elegans cultures may comprise microbes of a mammalian microbiome (e.g., a human microbiome). In some embodiments, one or more C. elegans cultures in such systems may comprise microbes of a canine, feline, equine, bovine, ovine, caprine, or porcine microbiome. The microbiomes used in accordance with the present disclosure may be obtained or derived from a targeted anatomical site of a mammalian subject. Examples of such microbiomes may include, but are not limited to, a skin microbiome, an oral microbiome, a nasal microbiome, a gastrointestinal microbiome, a brain microbiome, a lung microbiome, and / or a urogenital microbiome.

[0012] In some embodiments, the microorganisms in each C. elegans culture may comprise one or more microbial strains, hi some embodiments, the microorganisms in each culture may comprise a single microbial strain.

[0013] In some embodiments, one or more of the C. elegans cultures may constitute a therapeutic or nutritional supplement.

[0014] Also provided herein are methods for characterizing microbiomes using multiple C.elegans cultures. In some embodiments, the methods are for screening an individual's (e.g., mammalian, e.g., human) microbiome to determine whether a microbial strain or combination of microbial strains affects a mammalian disease or disorder. In some embodiments, the methods are for diagnosing an individual (e.g., mammalian, e.g., human) based on one or more microbial strains in the individual's microbiome. In some embodiments, the methods are for monitoring disease or pathology progression in an individual (e.g., mammalian, e.g., human) based on one or more microbial strains in the individual's microbiome.

[0015] In some embodiments described herein, a method is provided that includes adding microorganisms obtained from a mammalian microbiome to each of the C.elegans cultures of the system described herein. In some embodiments, the microorganisms in each C.elegans culture may constitute one or more microbial strains. In some embodiments, the microorganisms in each such culture may constitute a single microbial strain. The microbiome used in the methods described herein may be obtained or derived from a target anatomical site of a mammalian subject. Examples of such microbiomes may include, but are not limited to, a skin microbiome, an oral microbiome, a nasal microbiome, a gastrointestinal microbiome, a brain microbiome, a lung microbiome, and / or a urogenital microbiome.

[0016] In some embodiments, the method may include adding multiple microbial strains of a mammalian microbiome to multiple C. elegans cultures, where a different microbial strain is added to each C. elegans culture, where each culture comprises the same transgenic C. elegans strain, and where the transgenic C. elegans strain models a mammalian disease or condition.

[0017] In some embodiments, one or more of the C. elegans cultures may comprise a therapeutic agent or nutritional supplement. Thus, in some embodiments, the described methods may further comprise adding a therapeutic agent or nutritional supplement to one or more of the C. elegans cultures.

[0018] In some embodiments, the methods described herein may further include determining one or more parameter values ​​of the transgenic C. elegans strain in each of the C. elegans cultures. In some embodiments, such parameter(s) of the transgenic C. elegans are associated with the mammalian disease or condition that the transgenic C. elegans strain models. Exemplary such parameters of the transgenic C. elegans may include biological function or phenotype and / or level and / or activity of a molecule (e.g., a small molecule, protein, polypeptide, or transcript) associated with the mammalian disease or condition.

[0019] In some embodiments, the method may further comprise: (a) determining one or more parameter values ​​of a transgenic C. elegans strain in a C. elegans culture before adding the microbial strain to such culture; (b) determining the same one or more parameter values ​​of a transgenic C. elegans strain in such culture after adding the microbial strain to the C. elegans culture; and (c) comparing the one or more parameter values ​​determined before adding the microbial strain to the one or more parameter values ​​determined after adding the microbial strain.

[0020] In some embodiments, the techniques described herein can be used to characterize microbial strains of the human biome that are associated with human diseases or pathologies. Thus, in some such embodiments, the transgenic C. elegans strains involved in the systems and methods described herein model human diseases or pathologies. An exemplary human disease or pathology modeled by the transgenic C. elegans strain is Alzheimer's disease. In some embodiments, the disease or pathology (e.g., human disease or pathology) modeled by the transgenic C. elegans strain may include a disease or pathology associated with an altered or defective HIF-1 pathway (e.g., may include a cellular stress response). In some embodiments, for example, the disease or pathology modeled by the transgenic C. elegans strain may be an intraocular neovascular disease or disorder (e.g., diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma). In some embodiments, the disease or condition that the transgenic C. elegans strain models is diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma.

[0021] Thus, certain aspects described herein relate to techniques for characterizing microbial strains of the human biome. For example, one aspect provides a method comprising: (a) adding a microbial strain to a C.elegans culture, including a transgenic C.elegans strain that models Alzheimer's disease; and (b) determining whether the microbial strain affects one or more parameters of the transgenic C.elegans strain, where the one or more parameters are related to Alzheimer's disease. In some embodiments, the transgenic C.elegans strain may include a transgene encoding human ssApoE4 protein, human Aβ1-42 polypeptide, or human pseudophosphorylated tau protein.

[0022] In some embodiments, such methods further include (a) determining one or more parameter values ​​of a transgenic C. elegans strain in a C. elegans culture prior to adding the microbial strain to such culture, (b) determining the same one or more parameter values ​​of a transgenic C. elegans strain in such culture after adding the microbial strain to such culture, and (c) comparing the one or more parameter values ​​determined prior to adding the microbial strain to the one or more parameter values ​​determined after adding the microbial strain. Exemplary such one or more parameters include, but are not limited to, (i) a level of C. elegans paralysis, (ii) a level of amyloid plaques, (iii) a level of tau fibrils, (iv) a level of neuroinflammation, (v) a level of proteasome function, and / or (vi) a combination thereof.

[0023] In another aspect, the disclosure provides a method for characterizing a microbial strain of a human biome that is associated with an altered or defective HIF-1 pathway, comprising: (a) adding the microbial strain to a C. elegans culture, including a transgenic C. elegans strain that models a disease or condition associated with an altered or defective HIF-1 pathway, and (b) determining whether the microbial strain affects one or more parameters of the transgenic C. elegans strain. In some embodiments, such a transgenic C. elegans strain may include a transgene encoding the human prolyl hydroxylase EGLN, a human HIF transcription factor, or a human HIFα protein.

[0024] In some embodiments, such methods further include (a) determining one or more parameter values ​​of the transgenic C. elegans strain in culture before adding the microbial strain to the C. elegans culture; (b) determining the same one or more parameter values ​​of the transgenic C. elegans strain in culture after adding the microbial strain to the C. elegans culture; and (c) comparing the one or more parameter values ​​determined before adding the microbial strain to the one or more parameter values ​​determined after adding the microbial strain.

[0025] Exemplary such one or more parameters include, but are not limited to, (i) the level of neuroinflammation, (ii) the level of proteasome function, (iii) the level of C. elegans egg-laying rate, and / or (iv) combinations thereof.

[0026] Also within the scope of this disclosure are transgenic C. elegans strains expressing two or more of: (i) human ssApoE4, (ii) human Aβ1-42, (iii) human pseudophosphorylated tau, and (iv) UbV-GFP proteasome marker.

[0027] The present disclosure also describes, inter alia, the use of the provided transgenic C. elegans strains, systems, and / or methods to screen a mammalian microbiome for microbial strains that affect a mammalian disease or condition. Also included within the scope of the present disclosure is the use of the provided C. elegans, systems, and / or methods to characterize the impact that microbial strains of a mammalian microbiome have on a mammalian disease or condition. For example, a human microbiome can be screened / characterized using the techniques provided herein in accordance with the present disclosure.

[0028] The present disclosure describes, inter alia, compositions comprising one or more microbial strains. In some embodiments, compositions are provided herein that comprise one or more microbial strains, extracts thereof, and / or components thereof from a mammalian microbiome that have been assessed, identified, characterized, or assayed using transgenic C. elegans or methods as described herein. In some embodiments, compositions are provided herein that comprise 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, or 10 or more microbial strains, extracts thereof, and / or components thereof from a mammalian microbiome that have been assessed, identified, characterized, or assayed using transgenic C. elegans or methods as described herein.

[0029] In some embodiments, provided herein are compositions comprising 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, or 10 or more microbial strains listed in Table 8 below.

[0030] In some embodiments, provided herein are compositions comprising Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium, Bacillus subtilis, Acidaminococcus sp., or combinations thereof. In some embodiments, the combination includes at least two of, at least three of, at least four of, at least five of, at least six of, at least seven of, at least eight of, at least nine of, or all of Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium, Bacillus subtilis, and Acidaminococcus sp.

[0031] In some embodiments, the composition is a pharmaceutical composition. In some embodiments, the composition is an ingestible item.

[0032] The present disclosure describes, inter alia, methods that include administering the compositions described herein.

[0033] In some embodiments, a method of treating a disease or condition in a subject, comprising administering to a subject in need thereof a composition as described herein. In some embodiments, the disease or condition is a neurodegenerative disease or disorder. In some embodiments, the disease or condition is Alzheimer's disease. In some embodiments, the disease or condition may be associated with an altered or defective HIF-1 pathway. In some embodiments, the disease or condition may be an intraocular neovascular disease or disorder. In some embodiments, the disease or condition is diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma. The present disclosure describes, among other things, the use of the composition as described herein. In some embodiments, the use of the composition as described herein is in the treatment of a disease or condition in a subject. In some embodiments, the disease or condition is a neurodegenerative disease or disorder. In some embodiments, the disease or condition is Alzheimer's disease. In some embodiments, the disease or condition may be associated with an altered or defective HIF-1 pathway. In some embodiments, the disease or condition can be an intraocular neovascular disease or disorder, hi some embodiments, the disease or condition is diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma.

[0034] These and other aspects encompassed by the present disclosure are further detailed below and in the claims.

[0035] definition The scope of the present invention is defined by the claims appended hereto, and is not limited by any particular embodiment described herein. Upon review of this specification, one skilled in the art will recognize various modifications that may be equivalent to such described embodiments or may otherwise be within the scope of the claims. In general, the terms used herein follow their understood meanings in the art unless expressly indicated otherwise. Explicit definitions of certain terms are provided below, but the meaning of these and other terms in specific instances throughout this specification will be clear to those skilled in the art from the context.

[0036] The use of ordinal terms such as "first," "second," "third," etc. in the claims to modify claim elements does not, by itself, imply any priority, precedence, or ordering of one claim element over another claim element, or the temporal order in which acts of a method are performed, but rather is merely used as a label to distinguish one claim element having a particular name from another element having the same name (except for the use of ordinal terms) to distinguish the claim elements.

[0037] The articles "a" and "an" as used herein should be understood to include plural referents unless there is a clear indication to the contrary. A claim or description including "or" between one or more members of a group is deemed applicable if one, more than one, or all of the members of the group are present in, used in, or otherwise relevant to a given product or process, unless there is an indication to the contrary or otherwise clear from the context. In some embodiments, exactly one member of a group is present in, used in, or otherwise relevant to a given product or process. In some embodiments, more than one or all of the members of a group are present in, used in, or otherwise relevant to a given product or process. Unless otherwise indicated or unless a contradiction or inconsistency would be apparent to one skilled in the art, the present invention should be understood to encompass all variations, combinations, and permutations of one or more limitations, elements, clauses, descriptive terms, etc. from one or more of the enumerated claims that are introduced into another claim (or any other claim, if relevant) that is dependent on the same base claim. When elements are presented as a list (e.g., in a Markush group or similar format), it should be understood that each subgroup of the elements is also disclosed and any element(s) can be removed from the group. In general, when an embodiment or aspect is referred to as "comprising" certain elements, features, etc., it should be understood that a particular embodiment or aspect "consists of" or "consists essentially of" such elements, features, etc. For purposes of brevity, the embodiments are not expressly and specifically defined in this specification in every instance. It should also be understood that any embodiment or aspect may be expressly excluded from the claims, regardless of whether the specific exclusion is recited in the specification.

[0038] Administration: As used herein, the term "administration" typically refers to administration of a composition to a subject or system to achieve delivery of the agent to the subject or system. In some embodiments, the agent is a composition or is included in a composition. In some embodiments, the agent is produced through metabolism of the composition or one or more of its components. A person skilled in the art will recognize various routes that may be utilized for administration to a subject, e.g., a human, under appropriate circumstances. For example, in some embodiments, administration may be ocular, oral, parenteral, topical, etc. In some particular embodiments, administration may be bronchial (e.g., by bronchial instillation), buccal, dermal (which may be or may include one or more of, for example, topical to the dermis, intradermal, interdermal, transdermal, etc.), enteral, intra-arterial, intradermal, intragastric, intramedullary, intramuscular, intranasal, intraperitoneal, intrathecal, intravenous, intraventricular, in a specific organ (e.g., in the liver), mucosal, nasal, oral, rectal, subcutaneous, sublingual, topical, tracheal (e.g., by intratracheal instillation), vaginal, vitreous, etc. In many embodiments provided by the present disclosure, administration is oral administration. In some embodiments, administration may involve only a single dose. In some embodiments, administration may involve the application of a number of doses. In some embodiments, administration may involve intermittent (e.g., multiple doses spaced apart over time) and / or periodic (e.g., individual doses spaced apart by a common period of time) dosing. In some embodiments, administration may involve continuous dosing (e.g., perfusion) for at least a selected period of time. Administration of cells may be by any suitable route that results in delivery to a desired site in a subject, where at least a portion of the delivered cells or cell components remain viable. The period of viability of cells after administration to a subject may be as short as a few hours (e.g., 24 hours), to several days, or even as long as several years (i.e., long-term engraftment). In some embodiments, administration includes delivery of a bacterial extract or preparation that contains one or more metabolic products and / or by-products of the bacteria, but is completely devoid of viable bacterial cells.

[0039] Analog: As used herein, the term "analog" refers to a substance that shares one or more specific structural features, elements, components, or moieties with a reference substance. Typically, an "analog" exhibits significant structural similarity with a reference substance, for example, by sharing a core or consensus structure, but at the same time differs in a certain individual manner. In some embodiments, an analog is a substance that can be generated from a reference substance, for example, by chemical manipulation of the reference substance. In some embodiments, an analog is a substance that can be generated through the implementation of a synthetic process that is substantially similar to (e.g., shares multiple steps with) the synthetic process that generates the reference substance. In some embodiments, an analog is generated or can be generated through the implementation of a synthetic process that is different from the synthetic process used to generate the reference substance.

[0040] Approximately: When applied to one or more values ​​of interest, includes values ​​similar to a stated reference value. In certain embodiments, the term "approximately" or "about" refers to a range of values ​​that fall within ±10% (more or less) of a stated reference value, unless otherwise stated or otherwise clear from the context (except where such number exceeds 100% of possible values).

[0041] Equivalent: As used herein, the term "equivalent" refers to two or more agents, entities, states, sets of conditions, subjects, etc. that may not be identical to each other, but are sufficiently similar to permit a comparison between them, such that one of skill in the art will understand that a conclusion may be reasonably drawn based on the observed differences or similarities. In some embodiments, an equivalent set of conditions, circumstances, individuals, or populations is characterized by a number of substantially identical characteristics and one or a small number of variable characteristics. One of skill in the art will understand, in context, the degree of identity required for two or more such agents, entities, states, sets of conditions, etc. to be considered equivalent in any given situation. For example, one of skill in the art will understand that a set of circumstances, individuals, or populations is equivalent to each other when it is characterized by a sufficient number and type of substantially identical characteristics to warrant a reasonable conclusion that differences in results obtained or phenomena observed under or by the different sets of circumstances, individuals, or populations are caused by or indicative of variations in the variable characteristics.

[0042] Conservative: As used herein, refers to the case when describing conservative amino acid substitutions, which include replacing one amino acid residue with another amino acid residue having a side chain R group with similar chemical properties (e.g., charge or hydrophobicity). Generally, conservative amino acid substitutions do not substantially alter the desired functional properties of a protein, e.g., the ability of a receptor to bind a ligand. Examples of groups of amino acids having side chains with similar chemical properties include aliphatic side chains, such as glycine (Gly, G), alanine (Ala, A), valine (Val, V), leucine (Leu, L), and isoleucine (Ile, I); aliphatic-hydroxyl side chains, such as serine (Ser, S) and threonine (Thr, T); amide-containing side chains, such as asparagine (Asn, N) and glutamine (Gln, Q); aromatic side chains, such as phenylalanine (Phe, F), tyrosine (Tyr, Y), and tryptophan (Trp, W); basic side chains, such as lysine (Lys, K), arginine (Arg, R), and histidine (His, H); acidic side chains, such as aspartic acid (Asp, D) and glutamic acid (Glu, E); and sulfur-containing side chains, such as cysteine ​​(Cys, C) and methionine (Met, M). Conservative amino acid substitutions include, for example, valine / leucine / isoleucine (Val / Leu / Ile, V / L / I), phenylalanine / tyrosine (Phe / Tyr, F / Y), lysine / arginine (Lys / Arg, K / R), alanine / valine (Ala / Val, A / V), glutamic acid / aspartic acid (Glu / Asp, E / D), and asparagine / glutamine (Asn / Gln, N / Q). In some embodiments, a conservative amino acid substitution can be the substitution of any naturally occurring residue in a protein with alanine, for example, as used in alanine scanning mutagenesis. In some embodiments, conservative substitutions are made that have a positive value in the PAM250 log-likelihood matrix disclosed in Gonnet, GH et al., 1992, Science 256:1443-1445, which is incorporated herein by reference in its entirety. In some embodiments, the substitution is a moderately conservative substitution, and the substitution has a non-negative value in the PAM250 log-likelihood matrix.

[0043] [Table 1]

[0044] Control: As used herein, refers to the art-understood meaning of "control," which is a standard against which results are compared. Typically, controls are used to enhance the integrity of an experiment by isolating a variable in order to draw conclusions about such a variable. In some embodiments, a control is a reaction or assay that is performed simultaneously with a test reaction or assay to provide a comparison. "Control" also includes "control animals." A "control animal" may have a modification as described herein, a different modification than described herein, or no modification (i.e., a wild-type animal). In one experiment, the "test" (i.e., the variable being tested) is applied. In a second experiment, the "control," i.e., the variable being tested, is not applied. In some embodiments, a control is a historical control (i.e., of a previously performed test or assay, or of a known amount or result). In some embodiments, a control is or includes a printed or otherwise stored record. A control can be a positive or negative control.

[0045] Determine, measure, evaluate, assess, assay, and analyze: Determine, measure, evaluate, assess, assay, and analyze are used interchangeably herein to refer to any form of measurement, including determining whether an element is present or not. These terms include both quantitative and / or qualitative determinations. Assays can be relative or absolute. "Assaying for the presence of" can be determining the amount of something present and / or determining whether it is present or absent.

[0046] Dosage form: Those of skill in the art will appreciate that the term "dosage form" may be used to refer to a physically discrete unit of a drug (e.g., a therapeutic agent) for administration to a subject. Typically, each such unit contains a predetermined amount of drug. In some embodiments, such amount is a unit dosage amount (or all of it) appropriate for administration according to a dosing regimen (i.e., a therapeutic dosing regimen) that has been determined to correlate with a desired or beneficial outcome when administered to a relevant population. Those of skill in the art will appreciate that the total amount of a therapeutic composition or agent to be administered to a particular subject is determined by one or more attending physicians and may involve administration of multiple dosage forms.

[0047] Dosing regimen: Those skilled in the art will appreciate that the term "dosing regimen" may be used to refer to a set of unit doses (typically more than one) that are administered individually to a subject, typically spaced apart. In some embodiments, a given drug has a recommended dosing regimen, which may involve one or more doses. In some embodiments, a dosing regimen includes multiple doses, each of which is spaced apart in time from the other doses. In some embodiments, the individual doses are spaced apart from each other by the same length of time, and in some embodiments, a dosing regimen includes multiple doses, each of which is spaced apart by at least two different time periods. In some embodiments, all doses within a dosing regimen are of the same unit dose. In some embodiments, different doses within a dosing regimen are of different amounts. In some embodiments, a dosing regimen includes a first dose at a first dose, followed by one or more additional doses at a second dose that is different from the first dose. In some embodiments, the dosing regimen includes a first dose at a first dose, followed by one or more additional doses at a second dose that is the same as the first dose. In some embodiments, the dosing regimen correlates with a desired or beneficial outcome when administered across a relevant population.

[0048] Engineered: In general, the term "engineered" refers to an aspect of being manipulated by human beings. For example, a cell or organism is considered to be "engineered" if its genetic information has been manipulated to change (e.g., new genetic material not previously present has been introduced, for example, by transformation, mating, somatic hybridization, transfection, transduction, or other mechanisms, or previously present genetic material has been changed or removed, for example, by substitution or deletion mutations, or by mating protocols). As a convention and as will be understood by those skilled in the art, the progeny of an engineered polynucleotide or cell is typically still referred to as "engineered", even if the actual manipulation was performed on the prior entity.

[0049] Excipient: as used herein, refers to an inert (e.g., non-therapeutic) agent that can be included in a pharmaceutical composition, for example, to provide or aid in the desired consistency or stabilization effect. In some embodiments, suitable pharmaceutical excipients can include, for example, starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, skimmed milk powder, glycerol, propylene, glycol, water, ethanol, etc.

[0050] Functional: As used herein, a "functional" biomolecule is one that is in a form in which it exhibits a property and / or activity by which it is characterized. A biomolecule may have two functions (i.e., bifunctional) or many functions (i.e., multifunctional).

[0051] Gene: as used herein refers to a DNA sequence in a chromosome that codes for a product (e.g., an RNA product and / or a polypeptide product). In some embodiments, a gene includes coding sequences (i.e., sequences that code for a particular product). In some embodiments, a gene includes non-coding sequences. In some particular embodiments, a gene may include both coding (e.g., exon) and non-coding (e.g., intron) sequences. In some embodiments, a gene may include one or more regulatory sequences (e.g., promoters, enhancers, etc.) and / or intron sequences that may, for example, control or affect one or more aspects of gene expression (e.g., cell type specific expression, inducible expression, etc.). For the sake of clarity, it is noted that the term "gene" as used in this disclosure generally refers to a portion of a nucleic acid that codes for a polypeptide or a fragment thereof. As will be clear from the context to one of skill in the art, the term may optionally encompass regulatory sequences. This definition is not intended to exclude the application of the term "gene" to non-protein coding expression units, but is intended to clarify that the term as used herein most often refers to a polypeptide-encoding nucleic acid.

[0052] Improve, increase, enhance, inhibit, or reduce: As used herein, the terms "improve," "increase," "enhance," "inhibit," "reduce," or grammatical equivalents thereof, refer to a value that is relative to a baseline or other reference measurement. In some embodiments, the value is a statistically significant difference from the baseline or other reference measurement. In some embodiments, a suitable reference measurement may be or include a measurement in a particular system (e.g., in a single individual) under otherwise equivalent conditions except for (e.g., before and / or after) the presence of a particular agent or treatment, or in the presence of an appropriate equivalent reference agent. In some embodiments, a suitable reference measurement may be or include a measurement in an equivalent system that is known or expected to respond in a particular manner in the presence of the relevant agent or treatment. In some embodiments, a suitable reference is a negative reference, and in some embodiments, a suitable reference is a positive reference.

[0053] Isolated: As used herein, refers to a substance and / or entity that is (1) separated from at least some of the components with which it was associated when originally produced (whether in nature and / or in an experimental environment) and / or (2) designed, produced, prepared, and / or manufactured by human beings. In some embodiments, an isolated substance or entity may be concentrated, and in some embodiments, an isolated substance or entity may be pure. In some embodiments, an isolated substance and / or entity may be separated from about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% of the other components with which they were originally associated. In some embodiments, an isolated agent is about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% pure. As used herein, a substance is "pure" if it is substantially free of other components. In some embodiments, as will be understood by those of skill in the art, a substance may still be considered "enriched," "isolated," or even "pure" after being combined with certain other components, such as, for example, one or more carriers or excipients (e.g., buffers, solvents, water, etc.). In such embodiments, the isolation or percent purity of a substance is calculated without including such carriers or excipients. Those of skill in the art are aware of various techniques for isolating (e.g., concentrating or purifying) a substance or agent (e.g., using one or more of fractionation, extraction, precipitation, or other separations).

[0054] Pharmaceutical composition: As used herein, the term "pharmaceutical composition" refers to a composition in which an active agent is formulated together with one or more pharma- ceutically acceptable carriers. In some embodiments, the active agent is present in a unit dose suitable for administration in a treatment regimen that exhibits a statistically significant probability of achieving a predefined therapeutic effect when administered to a relevant population. In some embodiments, the pharmaceutical composition may be specially formulated for administration in solid or liquid form, including those adapted for oral administration, e.g., drenches (aqueous or non-aqueous liquids or suspensions), tablets, e.g., intended for buccal, sublingual, and systemic absorption, boluses, powders, granules, pastes for application to the tongue, capsules, powders, and the like. In some embodiments, the active agent may be or include a cell or cell population (e.g., a culture of, e.g., an EES microorganism). In some embodiments, the active agent may be or include an extract or component of a cell or cell population (e.g., a culture). In some embodiments, the active agent may be or include an isolated, purified, or pure compound. In some embodiments, the active agent may be synthesized in vitro (e.g., via chemical and / or enzymatic synthesis). In some embodiments, the active agent may be or include a natural product (whether isolated from its natural source or synthesized in vitro).

[0055] Pharmaceutically acceptable: As used herein, the term "pharmaceutical acceptable," which may be used, for example, in reference to a carrier, diluent, or excipient used to formulate a pharmaceutical composition as disclosed herein, means that the carrier, diluent, or excipient is compatible with the other ingredients of the composition and not deleterious to the recipient thereof.

[0056] Pharmaceutically acceptable carrier: As used herein, the term "pharmacologically acceptable carrier" means a pharma- ceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, excipient, or solvent encapsulating material, that is involved in carrying or transporting a compound of interest from one organ or body part to another. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient. Some examples of materials which can serve as pharma- ceutically acceptable carriers include sugars such as lactose, glucose, and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; glycols such as propylene glycol; polyols such as glycerin, sorbitol, mannitol, and polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; pH buffer solutions; polyesters, polycarbonates, and / or polyanhydrides; and other non-toxic, compatible substances used in pharmaceutical formulations.

[0057] Prevention: As used herein, the term "prevention" refers to delaying the onset of, and / or reducing the frequency and / or severity of, one or more symptoms of a particular disease, disorder, or condition. In some embodiments, prevention is assessed on a population basis, such that an agent is considered to "prevent" a disease, disorder, or condition if a statistically significant reduction in the onset, frequency, and / or intensity of one or more symptoms of the disease, disorder, or condition is observed in a population susceptible to the disease, disorder, or condition. In some embodiments, prevention may be considered complete, for example, if the onset of a disease, disorder, or condition is delayed for a given period of time.

[0058] Reference: as used herein describes a standard or control against which a comparison is made. For example, in some embodiments, an agent, animal, individual, population, sample, sequence, or value of interest is compared to a reference or control agent, animal, individual, population, sample, sequence, or value. In some embodiments, the reference or control is tested and / or determined substantially simultaneously with the test or determination of interest. In some embodiments, the reference or control is an existing reference or an existing control, optionally embodied in a tangible medium. Typically, as will be understood by those of skill in the art, the reference or control is determined or characterized under conditions or circumstances equivalent to those being assessed. Those of skill in the art will understand when there is sufficient similarity to indicate the reliability of and / or the validity of a comparison to a particular possible reference or control. In some embodiments, the reference is a negative control reference, and in some embodiments, the reference is a positive control reference.

[0059] Risk: As will be understood from the context, "risk" of a disease, disorder, and / or condition refers to the likelihood that a particular individual will develop the disease, disorder, and / or condition. In some embodiments, the risk is expressed as a percentage. In some embodiments, the risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, or up to 100%. In some embodiments, the risk is expressed as a relative risk to a risk associated with a reference sample or a group of reference samples. In some embodiments, the reference sample or a group of reference samples has a known risk of the disease, disorder, condition, and / or event. In some embodiments, the reference sample or a group of reference samples is derived from an individual comparable to the particular individual. In some embodiments, the relative risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more.

[0060] Sample: As used herein, the term "sample" typically refers to an aliquot of material obtained or derived from a source of interest. In some embodiments, the source of interest is a biological or environmental source. In some embodiments, the source of interest may be or include a cell or organism, such as a microorganism, a plant, or an animal (e.g., a human). In some embodiments, the source of interest is or includes a biological tissue or biological fluid. In some embodiments, the biological tissue or fluid may be or include amniotic fluid, aqueous humor, ascites, bile, bone marrow, blood, breast milk, cerebrospinal fluid, earwax, chyle, chyme, ejaculate, endolymph, exudate, stool, gastric acid, gastric juice, lymph, mucus, pericardial fluid, perilymph, peritoneal fluid, pleural fluid, pus, mucosal secretions, saliva, sebum, semen, serum, smegma, sputum, synovial fluid, sweat, tears, urine, vaginal fluid, vitreous humor, vomit, and / or combinations or component(s) thereof. In some embodiments, the biological fluid may be or include intracellular fluid, extracellular fluid, intravascular fluid (plasma), interstitial fluid, lymph, and / or transcellular fluid. In some embodiments, the biological fluid may be or include phytoexudates. In some embodiments, the biological tissue or sample may be obtained, for example, by aspiration, biopsy (e.g., fine needle or tissue biopsy), swab (e.g., oral, nasal, skin, or vaginal swab), scraping, surgery, washing or lavage (e.g., bronchoalveolar epithelium, duct, nasal, ocular, oral, uterine, vaginal, or other washing or lavage). In some embodiments, the biological sample is or comprises cells obtained from an individual. In some embodiments, the sample is a "primary sample" obtained directly from the intended source by any suitable means. In some embodiments, as will be clear from the context, the term "sample" refers to a preparation obtained by processing the primary sample (e.g., by removing one or more components of the primary sample and / or by adding one or more agents to the primary sample). For example, filtration using a semipermeable membrane.Such a "processed sample" may include, for example, nucleic acids or proteins extracted from a sample or obtained by subjecting a primary sample to one or more techniques, such as amplification or reverse transcription of nucleic acids, isolation and / or purification of certain components, etc.

[0061] Small molecule: As used herein, the term "small molecule" refers to a small organic or inorganic molecule having a molecular weight below about 3,000 Daltons. Generally, a small molecule may have a molecular weight less than 3,000 Daltons (Da). A small molecule can be, for example, at least about 100 Da to about 3,000 Da (e.g., about 100 to about 3,000 Da, about 100 to about 2500 Da, about 100 to about 2,000 Da, about 100 to about 1,750 Da, about 100 to about 1,500 Da, about 100 to about 1,250 Da, about 100 to about 1,000 Da, about 100 to about 750 Da, about 100 to about 500 Da, about 200 to about 1500, about 500 to about 1000, about 300 to about 1000 Da, or about 100 to about 250 Da).

[0062] Subject: As used herein, the term "subject" refers to an individual to whom the provided treatment is applied. In some embodiments, the subject is an animal. In some embodiments, the subject is a mammal, for example, a mammal experiencing or susceptible to a disease, disorder, or condition as described herein. In some embodiments, the animal is a vertebrate, for example, a mammal such as a non-human primate, (especially a higher primate), sheep, dog, rodent (e.g., mouse or rat), guinea pig, goat, pig, cat, rabbit, or cow. In some embodiments, the animal is a non-mammal, such as a chicken, amphibian, reptile, or invertebrate model C. elegans. In some embodiments, the subject is a human. In some embodiments, the patient is suffering from or susceptible to one or more diseases, disorders, or conditions as described herein. In some embodiments, the patient exhibits one or more symptoms of one or more diseases, disorders, or conditions as described herein. In some embodiments, the patient has been diagnosed with one or more diseases, disorders, or conditions as described herein. In some embodiments, the subject is undergoing or has undergone a particular therapy for diagnosing and / or treating a disease, disorder, or condition, hi another embodiment, the subject is an experimental animal or surrogate animal as a disease model.

[0063] Substantially: as used herein refers to a qualitative state of exhibiting the full or nearly full extent or degree of a desired property or characteristic. Those skilled in the art of biology will understand that biological and chemical phenomena rarely, if ever, proceed to completion and / or completeness, or achieve or avoid absolute results. The term "substantially" is therefore used herein to capture the potential lack of completeness inherent in many biological and chemical phenomena.

[0064] Therapeutic regimen: "Therapeutic regimen," as that term is used herein, refers to a dosing regimen, the administration of which across a relevant population can be correlated with a desired or beneficial therapeutic outcome.

[0065] Therapeutically effective amount: as used herein means an amount that produces the desired effect for which it is administered. In some embodiments, the term refers to an amount that is sufficient to treat a disease, disorder, and / or condition when administered to a population suffering from or susceptible to the disease, disorder, and / or condition according to a therapeutic dosing regimen. In some embodiments, a therapeutically effective amount is an amount that reduces the incidence and / or severity of and / or delays the onset of one or more symptoms of a disease, disorder, and / or condition. Those skilled in the art will appreciate that the term "therapeutically effective amount" does not actually require that successful treatment be achieved in a particular individual. Rather, a therapeutically effective amount may be an amount that, when administered to a patient in need of such treatment, provides a particular desired pharmacological response in a significant number of subjects. In some embodiments, reference to a therapeutically effective amount may be a reference to an amount as measured in one or more specific tissues (e.g., tissues affected by a disease, disorder, or condition) or fluids (e.g., blood, saliva, serum, sweat, tears, urine, etc.). One of ordinary skill in the art will appreciate that in some embodiments, a therapeutically effective amount of a particular agent or therapy may be formulated and / or administered in a single dose, hi some embodiments, a therapeutically effective agent may be formulated and / or administered in multiple doses, e.g., as part of a dosing regimen.

[0066] Treatment: As used herein, the term "treatment" (also "treat" or "treating") refers to any application of therapy that partially or completely alleviates, improves, relieves, inhibits, delays the onset of, reduces the severity of, and / or reduces the incidence of one or more symptoms, characteristics, and / or causes of a particular disease, disorder, and / or condition. In some embodiments, such treatment may be treatment of subjects who do not exhibit symptoms of the relevant disease, disorder, and / or condition and / or subjects who exhibit only early signs of the disease, disorder, and / or condition. Alternatively, or in addition, such treatment may be treatment of subjects who exhibit one or more established signs of the relevant disease, disorder, and / or condition. In some embodiments, treatment may be treatment of subjects who have been diagnosed as suffering from the relevant disease, disorder, and / or condition. In some embodiments, treatment may be treatment of subjects known to have one or more susceptibility factors that are statistically correlated with an increased risk of developing the relevant disease, disorder, and / or condition. [Brief description of the drawings]

[0067] [Figure 1] Data showing evidence that APOE4 protein augments Aβ-induced paralysis phenotype in C. elegans. C. elegans animals with the indicated genotypes were administered standard E. coli OP50 laboratory strain. Animals were monitored every other day from adulthood day 1 until all animals were paralyzed. At least 20 animals were scored for each assay. Data from three independent trials are presented in the plots. For each data point, the mean ± standard deviation is presented in the graph. [Diagram 2]Data are presented showing evidence that expression of Aβ3-42 conjugated to green fluorescent protein (GFP) and human ssAPOE4 significantly increased the number of aggregates. C. elegans animals with the indicated genotypes were administered standard E. coli OP50 laboratory strain. GFP aggregates in the anterior part of the animals were counted when the animals reached adulthood. At least 17 animals were scored for each assay. For each data point, the mean ± standard deviation is presented in the bar graph. Compared to animals expressing Aβ3-42, animals expressing Aβ3-42 and ssApoE4 had significantly increased GFP aggregates when analyzed using a Student's t-test (P<0.0001). [Diagram 3] Data are presented showing evidence that UbV-GFP is stabilized in animals expressing both human ssAPOE4 and human tau352 (PHP). C. elegans animals with the indicated genotypes were administered standard E. coli OP50 laboratory strain. When the animals reached adulthood, the number of animals expressing GFP in the intestine was counted. At least 30 animals were scored for each assay. Data from three independent trials are presented in bar graphs. For each data point, the mean ± standard deviation is presented on the graph. Compared to animals expressing tau352 (PHP), animals expressing ssAPOE4 and tau352 (PHP) had significantly increased UbV-GFP expression when analyzed using a Student's t test (P < 0.0001). [Figure 4] Data are included showing evidence that certain microbial strains of the microbiome increase Aβ-mediated paralysis. C. elegans animals with the indicated genotypes were administered either the standard E. coli OP50 laboratory strain or the individual microbiome strains. On adult day 4, the number of paralyzed animals was recorded. Data from three independent trials are presented in bar graphs. For each data point, the mean ± standard deviation is presented on the graph. Table 1 contains the raw data, including the number of animals analyzed for each condition. [Diagram 5]Data are included showing evidence that certain microbial strains of the microbiome modulate Aβ3-42::GFP aggregation. C. elegans animals with the indicated genotypes were administered either standard E. coli op50 laboratory strains or individual microbiome strains. When animals reached adulthood, the number of GFP aggregates in the anterior part of the animals was counted. GFP aggregates in three animals were counted for each condition. For each data point, the mean ± standard deviation is presented on the graph. [Figure 6] Contains data showing that certain microbial strains altered ATP production. Neuro2A cells were mock treated or treated with 108 CFU of each bacteria or a combination of all bacteria (CT10) for 16 hours. 2 μM human Abeta 1-42 was added to all wells except untreated wells. Cells were incubated for 24 hours and ATP levels were measured and normalized. ATP percentage was calculated according to the following formula: ATP% = [(normalized luminescence value of sample / normalized luminescence of control) x 100]. [Figure 7] 1 includes a schematic diagram of an exemplary HIF pathway. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0068] Provided herein are transgenic C. elegans disease models and methods of using the C. elegans disease models to rapidly identify, assess, or characterize one or more microbial strains from a mammalian microbiome that affect (e.g., enhance or minimize) a phenotype associated with a mammalian disease or pathology. In some embodiments, the identification, assessment, or characterization can occur with or without chemical entities and / or biological agents (e.g., antibodies).

[0069] C. elegans is a bacteriovorous nematode that is fed an E. coli diet in the laboratory. C. elegans can provide reliable, valid and efficient genotypic and phenotypic models for several mammalian diseases and conditions, including various human diseases and conditions such as aging, diabetes, neurodegenerative disorders, metabolic diseases, and cancer. The present disclosure provides insight that C. elegans models of mammalian diseases and conditions can be used to rapidly screen the human microbiome for microbial strains that affect such mammalian diseases and conditions. It is unlikely that such studies would be performed in standard mammalian cell culture assays or animal models. For example, screening millions of microbial species and / or strains individually, or even in combination, for example using mouse models of human diseases and conditions, would be economically unfeasible, time-consuming and laborious. Although C. elegans may not contain some of the complexities of mammalian systems, C. elegans models share characteristics with mammalian systems, are fast in response time, and are less expensive to produce and maintain than mammalian model systems. Thus, C. elegans models of mammalian disease and pathology provide a powerful first line of defense to examine the influence that microbial strains in the mammalian biome have on mammalian disease and pathology, and represent a useful tool to prioritize lead microbial strains that affect specific, sensitive and conserved therapeutic targets.

[0070] Provided herein is a method of using C.elegans as a tool for rapid screening of human microbiome for disease modifiers. Standard microbiology techniques can be used to culture microbial strains from samples from healthy patients or patients with disease or pathology. These microbial strains can be fed to transgenic C.elegans strains carrying mammalian (e.g., human) disease markers, mammalian (e.g., human) disease gene mutations, or combinations thereof. The microbial strains can be fed to transgenic C.elegans strains either individually or in combination. In some instances, individual microbial strains or combinations of microbial strains can be combined with chemical entities (e.g., small molecules, e.g., drugs) or biologics (e.g., monoclonal antibodies) and fed to transgenic C.elegans strains carrying mammalian (e.g., human) disease markers, mammalian (e.g., human) disease gene mutations, or combinations thereof.

[0071] The present disclosure recognizes that multiple outcomes may arise from feeding individual microbial strains or combinations of microbial strains in combination with chemical entities (e.g., small molecules, e.g., drugs) or biologics (e.g., monoclonal antibodies) to transgenic C. elegans strains carrying mammalian (e.g., human) disease markers, mammalian (e.g., human) disease gene mutations, or combinations thereof. In the first scenario, the methods described herein can be used to identify, define, assess, and / or detect individual microbial strains or combinations of microbial strains from a mammalian microbiome that increase the phenotypic severity of a mammalian disease or condition in C. elegans. In the second scenario, the methods described herein can be used to identify, define, assess, and / or detect individual microbial strains or combinations of microbial strains from a mammalian microbiome that decrease the phenotypic severity of a mammalian disease or condition in C. elegans. In a third scenario, the methods described herein can be used to identify, define, characterize, and / or detect individual microbial strains or combinations of microbial strains from a mammalian microbiome that do not have any phenotypic effects of a mammalian disease or condition in C. elegans. This disclosure recognizes that each of these outcomes provides valuable information.

[0072] For example, individual microbial strains or combinations of microbial strains from a mammalian microbiome that increase the phenotypic severity of a mammalian disease or condition in C. elegans may be potential early diagnostic biomarkers for a mammalian disease or condition. In some embodiments, such individual microbial strains or combinations of microbial strains correlate with a higher incidence or increased severity of a disease or condition in a mammal (e.g., human). Where such a correlation has not been found before, the transgenic C. elegans and methods using transgenic C. elegans described herein can be used to rapidly screen and / or evaluate one or more microbial strains in a mammalian microbiome using genetic screening or chemical extraction or genomic data mining methods to identify potential "toxic" metabolites or components of the microbiome that are responsible for the increased severity or incidence of the disease. These identified microbial strains and / or components of the mammalian microbiome can also be used to develop diagnostic methods. The identification and / or characterization of "toxic" microbial strains or microbiome components can also be used to develop modulators or therapeutic agents that can target the microbial strains, microbiome components, or the biosynthetic pathways that produce them.

[0073] In addition, the present disclosure recognizes that individual microbial strains or combinations of microbial strains from a mammalian microbiome that reduce the phenotypic severity of a mammalian disease or condition in C. elegans may correlate with the severity of the disease in a human patient. In some embodiments, such individual microbial strains or combinations of microbial strains correlate with a lower incidence or reduced severity of the disease or condition in a mammal (e.g., human). Where such a correlation has not been found before, the transgenic C. elegans and methods using transgenic C. elegans described herein can be used to rapidly screen and / or evaluate one or more microbial strains in a mammalian microbiome using genetic screening or chemical extraction or genomic data mining methods to identify potential "beneficial" microbial strains, microbiome components, or metabolites that are involved in reducing the severity or incidence of the disease. These identified microbial strains and / or components of the mammalian microbiome can also be used to develop diagnostic methods. The identification and / or characterization of "beneficial" microbial strains or microbiome components can also be used as modulators or therapeutics for the disease.

[0074] In some instances, individual microbial strains or combinations of microbial strains from a mammalian microbiome can also be fed to transgenic C. elegans strains in combination with chemical entities (e.g., small molecules, e.g., drugs) or biologics (e.g., monoclonal antibodies), or combinations thereof, to identify potential biological or signaling pathways or cellular target genes or pathways. Biological or signaling pathways or cellular target genes or pathways can include, but are not limited to, inflammation, insulin receptor, cell death, mitochondria, endoplasmic reticulum, proteasome, lipogenesis, and detoxification.

[0075] In some instances, individual or combinations of microbial strains from mammalian microbiomes can be combined with chemical entities (e.g., small molecules, e.g., drugs) or biological agents (e.g., monoclonal antibodies), or combinations thereof, to feed C.elegans strains to identify signaling pathways or sets (e.g., comprehensive sets) of target genes or pathways that can modulate or optimize the function of specific organelles relevant to a particular disease. For example, there are multiple pathways or targets that can be modulated to achieve optimal mitochondrial function, which is a key target of relevance in several neurodegenerative diseases, including Alzheimer's disease (AD). Biological targets for improving mitochondrial function include biogenesis, bioenergetics, hormesis, and / or repair. Using the transgenic C. elegans disease models and / or methods using C. elegans disease models described herein, it is possible to identify individual or combinatorial microbiome species and / or combinations with chemical entities (e.g., small molecules, e.g., drugs) or biologics (e.g., monoclonal antibodies), or combinations thereof, that can modify or improve or alter any of mitochondrial biogenesis, bioenergetics, hormesis, or repair, or combinations thereof. Thus, it is possible to identify and combine individual microbial strains or combinations of microbial strains from a mammalian microbiome, and / or chemical entities (e.g., small molecules, e.g., drugs) or biologics (e.g., monoclonal antibodies), or combinations thereof, that can target multiple pathways, for example to achieve optimal mitochondrial function.

[0076] C. elegans The free-living nematode C. elegans has been used extensively as a model system. C. elegans is inexpensive to culture, easy to physically manipulate, and has numerous genetic and molecular tools available for research. C. elegans is a simple multicellular organism, i.e., adults contain approximately 1,000 somatic cells, yet still have a variety of tissue types, such as muscle, nerve, and intestinal cells. C. elegans has a short generation time, which allows for rapid experimentation. C. elegans typically progresses from egg to larvae to fertile adults in a period of three days at room temperature. A single adult C. elegans can have 300-1,000 offspring, which allows for a significant number of animals to be used and then rapidly replenished in a relatively short time. Due to sexual dimorphism, C. elegans is useful for genetics. Self-fertilizing hermaphrodites can be maintained as homozygous mutants without the need for mating, and males can be used in genetic crosses. C. elegans are transparent throughout all stages of their life cycle, which provides the ability to see inside the organism. This allows for the observation of cellular events. It also allows for the use of phosphorescent, luminescent, and fluorescent reporters. RNA-mediated interference (RNAi) can also be used to manipulate protein expression in C. elegans, which may allow for rapid assessment of gene function. Another advantage of using the C. elegans model system is the ability to freeze and recover the animals, which allows for long-term storage.

[0077] C. elegans can be genetically modified using several techniques to produce transgenic C. elegans strains. The sexual dimorphism of C. elegans allows for relatively easy genetic manipulations to be performed following known procedures. For example, if a strain needs to be propagated, a single hermaphrodite can be used to self-fertilize and produce a population of offspring. Even if an animal is unable to mate due to a mutation, the hermaphrodite can still produce offspring. Another aspect of C. elegans reproduction that makes it a useful genetic tool is the ability of the animal to mate with a male hermaphrodite. For example, mating experiments allow genetic markers, such as mutations that cause visible phenotypes, to be placed together in a single organism with unknown mutations to facilitate mapping of the mutations. Hermaphrodites produce only a limited number of sperm and can typically have approximately 300 self-offspring. Mating increases the number of offspring produced by a single hermaphrodite to approximately 1,000 due to the addition of sperm produced by the male. The relatively large number of progeny combined with the short life span of C. elegans allows rapid and inexpensive assays to be performed on the animals.

[0078] In addition to genetic modification via reproduction, C. elegans can be genetically modified via injection of transgenes. Microinjection is an effective method for producing transgenic animals and for directly introducing various types of molecules into cells. With regard to DNA transformation, one approach is to inject DNA into the distal arm of the gonad of C. elegans. The distal germline of C. elegans contains a central core of cytoplasm shared by many germ cell nuclei. Thus, DNA injected into the distal arm of the gonad of C. elegans can be delivered to many offspring. Direct microinjection into the nucleus of an oocyte can induce integration of the transgene into the chromosome, but this technique can be more difficult to perform. C. elegans can also integrate genetic material that is fed to them.

[0079] C. elegans is relatively simple to culture. It can be cultured in the presence of bacteria, either in liquid culture or on nematode growth medium (NGM) agar plates. It is possible to grow the animals in a chemically defined medium without the addition of bacteria, which can be useful because the components of the medium can be varied to study the nutritional or other chemical requirements of the animals. In some embodiments, C. elegans is grown on agar plates. C. elegans can be grown on nematode growth medium (NGM) agar plates. Bacteria can be spread on NGM plates as a food source for the animals. For example, OP50, a leaky E. coli uracil auxotroph, can be used. OP50 grows slowly and provides nutrients to the animals without overgrowing them. Once the animals have eaten all of the food on the plate, they burrow into the agar and can be maintained on "starvation" plates in a 15°C incubator for weeks at a time. The animals can be transferred to agar plates with fresh bacteria, either by cutting and removing a small block of agar from the starvation plate with a sterile instrument such as a micropipette tip, or by washing the worms off the surface of the plate with sterile water, or by picking one or more individuals onto a fresh plate, which will result in the re-emergence of C. elegans. At any point, C. elegans can be cryogenically preserved. C. elegans prefers to grow at 15°C to 25°C, but temperatures can vary depending on the strain of C. elegans and the conditions being tested. In some embodiments, C. elegans cultures can be cultured at temperatures of at least 5°C, at least 10°C, at least 15°C, at least 20°C, at least 25°C, at least 30°C, at least 35°C, or at least 40°C. In some embodiments, C. elegans cultures may be cultured at temperatures of at most 65°C, at most 60°C, at most 55°C, at most 50°C, at most 55°C, at most 40°C, at most 35°C, at most 30°C, at most 25°C, or at most 20°C.Standard protocols for C. elegans manipulation and culture are known, for example as described in Stiernagle T. Maintenance of C. elegans. Wormbook, ed. The C. elegans Research Community, WormBook. (February 11, 2006), which is incorporated herein by reference.

[0080] Microbial preparation(s) and / or component(s) The present disclosure provides systems and methods for assessing, characterizing, and identifying one or more microbial strains of a microbiome. Such systems and methods may be useful for assessing, characterizing, and identifying one or more microbial strains that affect the health of humans, livestock, and / or pets. In some embodiments, assessing, characterizing, and identifying one or more microbial strains from a snake, lizard, fish, or bird microbiome. In some embodiments, assessing, characterizing, and identifying one or more microbial strains from a mammalian microbiome. The mammalian microbiome may be a canine, feline, equine, bovine, ovine, caprine, or porcine microbiome. In general, the microbiome used in the systems or methods described herein will correspond to the disease or condition that the transgenic C. elegans used in the systems or methods is modeling. For example, if the transgenic C. elegans is modeling a human disease, then the human microbiome will be assessed, characterized, or identified.

[0081] The microbiome can be isolated from any system or tissue of an organism that supports microbial growth. For example, the microbiome can be a skin microbiome, an oral microbiome, a nasal microbiome, a gastrointestinal microbiome, a brain microbiome, a lung microbiome, or a urogenital microbiome. A list of exemplary microbial strains found in the gastrointestinal microbiome is included below in Table 8. Those skilled in the art will appreciate that microbiome samples can be obtained in a variety of ways known in the art. For example, skin, oral, nasal, lung, or urogenital microbiome samples can be obtained using swabs or tissue scrapes. In some embodiments, the gastrointestinal microbiome can be sampled from stool. Skin microbiome, oral microbiome, nasal microbiome, gastrointestinal microbiome, brain microbiome, lung microbiome, or urogenital microbiome samples can be obtained via biopsy.

[0082] In some embodiments, the microbiome is the microbiome of a healthy individual or an individual who does not have or is not at risk for a particular disease or disorder. In some embodiments, the microbiome is the microbiome of an individual who has or is at risk for a particular disease or disorder. In some embodiments, the microbiome is the microbiome of an individual who is known to have a particular disease or disorder. In some embodiments, the human microbiome is the microbiome of a human who is not known to be at risk for one or more diseases or conditions.

[0083] In some embodiments, the microbiome is a reference microbiome. The reference microbiome can be the microbiome of a healthy individual or an individual who is not suffering from or at risk of developing a particular disease or disorder. In some cases, the reference microbiome may be derived from the same individual as the microbiome to be assessed or characterized, but obtained at a different time. In some cases, the reference microbiome may be derived from the same individual as the microbiome to be assessed or characterized, but obtained from a different system or tissue.

[0084] In some embodiments, individual microbial strains or combinations of microbial strains may be assessed, characterized, or identified in relative abundances that are different from the relative abundances that such strain(s) are found in the microbiome.For example, the transgenic C. elegans or methods of using transgenic C. elegans described herein may be used to assess, characterize, or identify a single strain, even if it naturally exists in the microbiome with other microbial strains.As another example, the transgenic C. elegans or methods of using transgenic C. elegans described herein may be used to assess, characterize, or identify two microbial strains together, even if it naturally exists in the microbiome with additional microbial strains.

[0085] Also, the transgenic C. elegans or the method of using transgenic C. elegans described herein may be used to evaluate, characterize, or identify the extracts, components, or compounds of microbial strains. In some instances, the extracts, components, or compounds of microbial strains that are determined to affect transgenic C. elegans models of disease or pathology may be evaluated, characterized, or identified. The evaluation, characterization, or identification of the extracts, components, or compounds of microbial strains that affect transgenic C. elegans models of disease or pathology may provide additional information about potential biomarkers, targets, or protective factors in the microbiome.

[0086] A variety of techniques are known in the art that can be used to prepare extracts of microbial strains and / or to isolate or process extracts, components, or compounds from microbial strains (e.g., to isolate and / or purify one or more components or compounds therefrom). Such techniques may include, for example, one or more of organic extraction, vacuum concentration, chromatography, etc., to name just a few examples.

[0087] Biological effects assessment The present disclosure provides insight that C. elegans can be used to identify, characterize, or assess microbial strain(s) of a mammalian microbiome by contacting (e.g., feeding, administering) the microbial strain(s) to transgenic C. elegans that models a mammalian disease or condition. To determine whether a microbial strain or combination of microbial strains affects a transgenic C. elegans that models a mammalian disease or condition, parameters of the transgenic C. elegans can be observed, measured, or assessed in different samples that are contacted with the microbial strain or combination of microbial strains. To determine whether a microbial strain or combination of microbial strains affects a transgenic C. elegans that models a mammalian disease or condition, parameters of the transgenic C. elegans can be observed, measured, or assessed. As just a few examples, the behavior of transgenic C. elegans (e.g., mating, feeding, food aversion, or movement), genetic mutations (e.g., the presence of SNPs, deletions, additions, inversions, or repeats in DNA), transcript levels, protein levels, metabolite levels, lipid levels, carbohydrate levels, protein (e.g., enzyme) activity levels can be observed, measured, or assessed to determine whether a microbial strain or combination of microbial strains affects transgenic C. elegans that models a mammalian disease or condition.

[0088] In some embodiments, the methods described herein utilize a first sample and a second sample. In some embodiments, the first sample is a reference sample. In some embodiments, the reference sample can be a culture of transgenic C.elegans that has been contacted (e.g., administered or fed) with, for example, OP50. In some embodiments, the reference sample can be a culture of transgenic C.elegans that has been contacted (e.g., administered or fed) with a microbial strain or combination of microbial strains from the microbiome of a healthy individual. In some embodiments, the reference sample can be a culture of transgenic C.elegans that has been contacted (e.g., administered or fed) with a microbial strain or combination of microbial strains from the microbiome of an individual obtained at a first time point.

[0089] In some embodiments, the second sample can be a test sample. In some embodiments, the test sample can be a culture of transgenic C.elegans that has been contacted (e.g., administered or fed) with an individual or combination of microbial strains from a mammalian microbiome, such as a human microbiome. In some cases, the human microbiome is the microbiome of a human suffering from or at risk of a disease or condition. In some cases, the human microbiome is the microbiome of a human not known to be at risk of one or more diseases or conditions. In some embodiments, the test sample can be a culture of transgenic C.elegans that has been contacted (e.g., administered or fed) with an individual or combination of microbial strains from a microbiome of an individual obtained at a second time point.

[0090] In some embodiments, the methods described herein include comparing one or more parameters obtained from a test sample with one or more parameters obtained from a reference sample. In some embodiments, by comparing one or more parameters obtained from a test sample with one or more parameters obtained from a reference sample, it can be determined that an individual microbial strain or combination of microbial strains from a microbiome increases the severity or incidence of a phenotype of a disease or condition modeled by cultured transgenic C. elegans. In some embodiments, by comparing one or more parameters obtained from a test sample with one or more parameters obtained from a reference sample, it can be determined that an individual microbial strain or combination of microbial strains from a microbiome decreases the severity or incidence of a phenotype of a disease or condition modeled by cultured transgenic C. elegans. In some embodiments, by comparing one or more parameters obtained from a test sample with one or more parameters obtained from a reference sample, it can be determined that an individual microbial strain or combination of microbial strains from a microbiome has no effect on the severity or incidence of a phenotype of a disease or condition modeled by cultured transgenic C. elegans.

[0091] The transgenic C. elegans and methods of using transgenic C. elegans provided herein may be useful in assessing, characterizing, or identifying microbial strains of the microbiome that affect a mammalian disease or condition. The present disclosure also provides the recognition that the transgenic C. elegans and methods of using transgenic C. elegans provided herein may be used to define and / or characterize a microbial signature associated with a disease or condition. Furthermore, the present disclosure provides the recognition that the transgenic C. elegans and methods of using transgenic C. elegans provided herein may be used to define and / or characterize a microbial signature associated with one or more features of a disease or condition (e.g., severity, responsiveness to therapy, etc.). For example, when multiple microbial strains are determined to be associated with a disease or disorder of increased severity, for example across multiple individuals, the microbial strains, as well as their relative abundance, may be used as a signature to identify individuals at risk of developing a disease or disorder of increased severity. As another example, if multiple microbial strains are determined to be associated with increased severity of a disease or disorder, e.g., in a single individual, at a particular time point (e.g., after withdrawal from treatment), the microbial strains, as well as their relative abundance, can be used as a signature to identify when the individual is at risk of developing a disease or disorder of increased severity.

[0092] The present disclosure also provides the recognition that the transgenic C. elegans and methods of using transgenic C. elegans provided herein can be used to diagnose individuals having a disease or condition. Indeed, microbial signatures associated with a disease or condition determined through the use of the transgenic C. elegans and methods of using transgenic C. elegans provided herein can be used to provide an early diagnosis and / or identify individuals as at risk.

[0093] The present disclosure also provides the recognition that the transgenic C. elegans and methods of using the transgenic C. elegans provided herein can be used to monitor the progression of a disease or condition in an individual. For example, if the relative abundance within the microbiome of a microbial strain determined to increase the severity of a disease or condition decreases, this may indicate that the disease or condition is being alleviated, for example, by a treatment or immune response.

[0094] The present disclosure also provides insight that the transgenic C. elegans and methods of using transgenic C. elegans provided herein can be used to tailor treatments (e.g., therapies, nutritional supplements, and / or probiotics) to individual patients. In some embodiments, the transgenic C. elegans and methods of using transgenic C. elegans provided herein can provide "individualized" therapy. In some instances, microbial strains within an individual can be assessed, characterized, or identified to determine whether they have an impact on a disease or disorder. Based on the results, the individual can be treated with one or more microbial strains to adjust the microbial strains (and / or their components or compounds) in their microbiome. In some cases, this will have an impact on the disease or condition that the individual suffers from or is at risk of developing. For example, if an individual is determined to have a relatively low amount of one or more microbial strains determined to reduce the severity of a disease or condition, administration of the one or more microbial strains determined to reduce the severity of a disease or condition to that individual (or an extract, component, or compound thereof) may ameliorate the severity of the disease or condition in that individual.

[0095] Pharmaceutical Compositions Compositions comprising individual microbial strains or combinations of microbial strains are provided herein. In some embodiments, the compositions comprise individual microbial strains or combinations of microbial strains from a mammalian microbiome, extracts thereof, and / or components thereof, assessed, identified, characterized, or assayed using transgenic C. elegans or methods as described herein. In some embodiments, compositions are provided herein comprising 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, or 10 or more microbial strains from a mammalian microbiome, extracts thereof, and / or components thereof, assessed, identified, characterized, or assayed using transgenic C. elegans or methods as described herein.

[0096] In some embodiments, provided herein are compositions comprising 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, or 10 or more microbial strains listed in Table 8 below.

[0097] In some embodiments, provided herein are compositions comprising Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium, Bacillus subtilis, Acidaminococcus sp., or combinations thereof. In some embodiments, the combination includes at least two of, at least three of, at least four of, at least five of, at least six of, at least seven of, at least eight of, at least nine of, or all of Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium, Bacillus subtilis, and Acidaminococcus sp.

[0098] In some embodiments, individual or combination of microbial strains from a mammalian microbiome are killed (e.g., heat killed). Alternatively, in some embodiments, individual or combination of microbial strains from a mammalian microbiome may include viable or living cells.

[0099] In some embodiments, the one or more microbial strains include viable or living individual microbial strains or combinations of microbial strains, e.g., from a mammalian microbiome.

[0100] In some embodiments, the one or more microbial strains include viable or living individual microbial strains or combinations of microbial strains, e.g., from a mammalian microbiome, as described herein, and include and / or are formulated through the use of one or more cell cultures and / or supernatants or pellets thereof, and / or powders formed therefrom.

[0101] In some embodiments, a composition for use according to the present disclosure is a pharmaceutical composition, e.g., for administration (e.g., oral administration) to a mammal (e.g., a human). A pharmaceutical composition typically includes an active agent (e.g., an individual microbial strain or combination of microbial strains from a mammalian microbiome, an extract thereof, and / or components thereof) and a pharmaceutically acceptable carrier. Certain exemplary pharmaceutically acceptable carriers include, for example, saline, solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc., that are compatible with pharmaceutical administration.

[0102] In some embodiments, pharmaceutical compositions for use in accordance with the present disclosure may contain and / or be administered in combination with one or more supplementary active compounds. In certain embodiments, such supplemental active agents include ginger, curcumin, probiotics (e.g., probiotic strains of one or more of the following genera: Lactobacillus, Bifidobacterium, Saccharomyces, Enterococcus, Streptococcus, Pediococcus, Leuconostoc, Bacillus, and / or Escherichia coli (see Fijan, Int J Environ Res Public Health. 2014 May;11(5):4745-4767, which is incorporated herein by reference); prebiotics (non-digestible food components that help support the growth of probiotic bacteria, e.g., fructans such as fructooligosaccharides (FOS) and inulin, galactans such as galactooligosaccharides (GOS), dietary fibers such as resistant starch, pectin, beta-glucan, and xylooligosaccharides (Hutkins et al., Curr Opin Biotechnol. 2016 Feb;37:1-7, which is incorporated herein by reference), as well as combinations thereof.

[0103] A pharmaceutical composition is typically formulated to be compatible with its intended route of administration. An example of an administration route includes oral administration. Methods for formulating suitable pharmaceutical compositions are known in the art, see, for example, Remington: The Science and Practice of Pharmacy, 21st ed., 2005, and a series of books, Drugs and the Pharmaceutical Sciences: a Series of Textbooks and Monographs (Dekker, NY). Oral compositions generally include an inert diluent or an edible carrier. In some embodiments, oral formulations can be or include syrups, liquids, tablets, lozenges, gummies, capsules, such as gelatin capsules, powders, gels, films, and the like, to name just a few examples.

[0104] In some embodiments, pharma- ceutical compatible binders and / or adjuvant materials may be included as part of the pharmaceutical composition. In some particular embodiments, the pharmaceutical composition may contain, for example, any one or more of the following inactive ingredients or compounds of similar nature: binders such as microcrystalline cellulose, tragacanth gum, or gelatin; excipients such as starch or lactose, disintegrants such as alginic acid, Primogel, or corn starch; lubricants such as magnesium stearate or Sterotes; glidants such as colloidal silicon dioxide; sweeteners such as sucrose or saccharin; or flavorings such as peppermint, methyl salicylate, or orange flavor. In some embodiments, the composition may be consumed as is, or sprinkled or mixed into food or liquid (such as water). In some embodiments, a composition that may be administered to a mammal as described herein may be or may comprise an ingestible item (e.g., a food or beverage) that contains (e.g., is supplemented with) individual microbial strains or combinations of microbial strains from the mammalian microbiome, extracts thereof, and / or components thereof.

[0105] In some embodiments, the food product may be or include one or more of bars, candies, baked goods, cereals, savory snacks, pasta, chocolate, and other solid foods, liquid or semi-solid foods including yogurt, soups, and stews, and beverages such as smoothies, shakes, juices, and other carbonated or non-carbonated beverages. In some embodiments, the food product is prepared by the subject by mixing in individual or combinations of microbial strains from a mammalian microbiome, extracts thereof, and / or components thereof.

[0106] The compositions can be included in a kit, container, pack, or dispenser together with instructions for administration or use in the methods described herein.

[0107] Upon review of this disclosure, one of skill in the art will understand that in some embodiments, a composition (e.g., a pharmaceutical composition) as described herein may be or may include one or more cells, tissues, or organisms (e.g., plant or microbial cells, tissues, or organisms) that produce (e.g., have produced and / or are producing) the relevant compound.

[0108] Those skilled in the art will appreciate that in some embodiments, techniques for preparing and / or preparing compositions and / or preparations (and particularly for preparing pharmaceutical compositions) may include one or more steps of assessing or characterizing the compound, preparation, or composition, e.g., as part of quality control. In some embodiments, if the assayed material does not meet predefined specifications for the relevant assessment, it is discarded. In some embodiments, if such assayed material does in fact meet predefined specifications, it continues to be processed as described herein.

[0109] In some embodiments, the pharmaceutical compositions provided herein may promote colonization of individual microbial strains or combinations of microbial strains from a mammalian microbiome, particularly microbial strain(s) that have been identified, characterized, or assessed to reduce the severity or incidence of a mammalian disease or condition in a mammal suffering from or at risk for the mammalian disease or condition. In some embodiments, the pharmaceutical compositions provided herein may attenuate colonization of individual microbial strains or combinations of microbial strains from a mammalian microbiome, particularly microbial strain(s) that have been identified, characterized, or assessed to increase the severity or incidence of a mammalian disease or condition in a mammal suffering from or at risk for the mammalian disease or condition. In some embodiments, the pharmaceutical compositions provided herein may promote colonization of individual microbial strains or combinations of microbial strains from a mammalian microbiome, in particular microbial strain(s) that have been identified, characterized, or assessed as not affecting the severity or incidence of a mammalian disease or condition, but that are capable of outcompeting one or more microbial strains that have been identified, characterized, or assessed as increasing the severity or incidence of a mammalian disease or condition in a mammal suffering from or at risk for a mammalian disease or condition.

[0110] In some embodiments, each of the one or more microbial strains in the composition is at least 10 1 ~10 12 In some embodiments, each of the one or more microbial strains in the composition comprises 10 6 ~10 12 In some embodiments, each of the one or more microbial strains in the composition comprises the same number of CFU. In some embodiments, some of the one or more microbial strains in the composition comprise different numbers of CFU.

[0111] In some embodiments, the composition comprises a total of 10 6 ~10 12 Contains CFU.

[0112] In some embodiments, a pharmaceutical composition is tailored to a particular mammal (e.g., a particular human patient) based on the microbiome of that particular mammal (e.g., human). In some embodiments, a pharmaceutical composition is specific to the microbiome of an individual mammal (e.g., human). In some embodiments, a pharmaceutical composition is specific to the microbiome of a population of mammals (e.g., humans). A population of mammals can include, but is not limited to, a family, mammals in the same geographic location (e.g., neighborhood, city, state, or country), mammals having the same disease or condition, mammals of a particular age or age range, mammals consuming a particular diet (e.g., food, food source, or caloric intake).

[0113] Treatment The present disclosure recognizes that the compositions described herein may be useful in treating a subject. The methods provided by the present disclosure include methods for the treatment of certain diseases, disorders, and conditions. In some embodiments, the relevant disease, disorder, and condition may be or include a neurodegenerative disease, disorder, or condition. In some embodiments, the neurodegenerative disease, disorder, or condition may be Alzheimer's disease. In some embodiments, the relevant disease, disorder, and condition may be or include an intraocular neovascular disease, disorder, or condition. In some embodiments, the neurodegenerative disease, disorder, or condition may be diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma.

[0114] In general, the methods of treatment provided by the present disclosure involve administering a therapeutically effective amount of a composition as described herein, alone or in combination with other compositions and / or treatments, to a subject in need of, or determined to be in need of, such treatment.

[0115] In some embodiments, the methods of treatment provided herein are prophylactic or preventative, e.g., may be administered to a subject prior to the onset of noticeable symptoms and / or prior to exposure to a particular expected trigger associated with a neurodegenerative disease, disorder, or condition. In some embodiments, the methods of treatment provided herein are therapeutic, e.g., may be administered to a subject after the onset of noticeable symptoms associated with a neurodegenerative disease, disorder, or condition.

[0116] In some embodiments, the provided methods of treatment are administered to a subject that is a mammal, e.g., a mammal experiencing a disease, disorder, or condition as described herein, hi some embodiments, the subject is a human subject or a non-human veterinary subject, e.g., an ape, cat, dog, monkey, or pig.

[0117] In many embodiments, treatment involves ameliorating at least one symptom of a disease, disorder, or condition associated with a neurodegenerative disease, disorder, or condition. In some embodiments, the method of treatment may be prophylactic.

[0118] In some embodiments, the methods may involve administration of a therapeutically effective amount of a composition disclosed herein prior to, during (e.g., simultaneously with), or after administration of a treatment suspected to be associated with a neurodegenerative disease, disorder, or condition.

[0119] In some embodiments, a subject undergoing treatment as described herein may be undergoing and / or may have undergone other treatments (e.g., drug treatments / therapies, surgery, etc.) that may be intended to treat, for example, one or more symptoms or features of a disease, disorder, or condition as described herein (e.g., a neurodegenerative disease, disorder, or condition), whereby the provided compositions are administered in combination with such other therapies (i.e., treatments) to treat the relevant disease, disorder, or condition.

[0120] In some embodiments, the compositions described herein may be administered in a form containing one or more pharma- ceutically acceptable carriers. Suitable carriers have been described previously and vary depending on the desired form and mode of administration of the composition. For example, pharma- ceutically acceptable carriers may include diluents or excipients, such as fillers, binders, wetting agents, disintegrants, surfactants, glidants, and lubricants. Typically, the carriers may be solid (including powders), liquid, or any combination thereof. Each carrier is preferably "acceptable" in the sense of being compatible with the other ingredients in the composition and not harmful to the subject. The carrier may be biologically acceptable and inert (e.g., it allows the composition to maintain the viability of the biomaterial until it is delivered to the appropriate site).

[0121] The tablets, pills, capsules, troches and the like may contain any of the following ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth, or gelatin; an excipient such as starch or lactose, a disintegrating agent such as alginic acid, primogel, or corn starch; a lubricant such as magnesium stearate or sterote; a glidant such as colloidal silicon dioxide; a sweetening agent such as sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, orange flavor, or other suitable flavor. These are by way of example only and are not intended to be limiting.

[0122] Oral compositions may include inert diluents or edible carriers. For the purpose of oral therapeutic administration, the active compound may be incorporated with excipients and used in the form of tablets, lozenges, lozenges, troches, or capsules, such as gelatin capsules. Oral compositions may also be prepared by combining the compositions of the present disclosure with food. In some embodiments, the microorganisms may be formulated in food products. Some non-limiting examples of food products that may be used with the methods and compositions described herein include popsicles, cheese, cream, chocolate, dairy, meat, beverages, spices, kefir, miso, sauerkraut, and the like. In other embodiments, the foodstuffs may be juices, soft drinks, tea-based drinks, drink preparations, jelly drinks, and functional drinks; alcoholic drinks such as beer; carbohydrate-containing foods such as rice processed foods, noodles, bread, and pasta; fish, ham, sausage, fish paste products, and other paste products; retort pouch products such as curry, foods with thick sauces, and Chinese soup; soup; dairy products such as milk, milk drinks, ice cream, and yogurt; fermented products such as miso, fermented drinks, and pickles; soy products; various confectionery products including biscuits, cookies, candy, chewing gum, gummies, cold desserts including jellies, custard puddings, and frozen desserts; instant foods such as instant soups and instant miso soups; and the like. It is preferred that the food preparation does not require cooking after mixing with the microbial strain(s) to avoid the death of any microorganisms. In one embodiment, the food used for administration is chilled, for example ice-cold flavored water. In certain embodiments, the foodstuff is not a potentially allergenic foodstuff (e.g., not soy, wheat, peanut, tree nut, dairy, egg, shellfish, or fish). Pharmaceutically compatible binding agents, and / or adjuvant materials can be included as part of the composition.

[0123] In some such embodiments, the compositions described herein are administered to a subject according to a dosing regimen that achieves population of the subject's microbiome with the administered cells. In some embodiments, the compositions are administered to a subject in a single dose. In some embodiments, the compositions are administered to a subject in multiple doses. In some embodiments, doses of the compositions are administered to a subject twice a day, daily, weekly, or monthly.

[0124] In some embodiments, each of the one or more microbial strains in a dose is at least 10 1 ~10 12 In some embodiments, each of the one or more microbial strains in a dose comprises 10 6 ~10 12 In some embodiments, each of the one or more microbial strains in a dose contains the same number of CFU. In some embodiments, some of the one or more microbial strains in a dose contain different numbers of CFU.

[0125] In some embodiments, a dose of one or more microbial strains comprises a total of 10 6 ~10 12 In some embodiments, a dose of one or more microbial strains comprises a total of 10 7 ~10 10 In some embodiments, a dose of one or more microbial strains comprises 5-200 billion CFU. In some embodiments, a dose of one or more microbial strains comprises 5-50 billion CFU. In some embodiments, a dose of one or more microbial strains comprises 5-20 billion CFU. In some embodiments, a dose of one or more microbial strains comprises 50-100 billion CFU. In some embodiments, a dose of one or more microbial strains comprises 100-200 billion CFU. EXAMPLES

[0126] The following examples are provided to illustrate to one of ordinary skill in the art how to make and use the methods and compositions described herein and are not intended to limit the scope of the disclosure.

[0127] Example 1: Materials and Methods Two different constructs for the human APOE4 transgene under the control of the intestinal promoter (pvha-6::apoE4::tbb-2 UTR) were generated by gene synthesis. The constructs were cloned into the KpnI / SalI restriction enzyme sites of the pUC57 plasmid. The human APOE4 protein sequence was codon-optimized for optimal expression in C. elegans. Three synthetic introns were included within the apoE4 sequence in both constructs to avoid gene silencing and for optimal expression in C. elegans. The nucleotide sequences of the introns are included in Table 1 below.

[0128] In one construct, the signal sequence FLP-1 for secretion from C. elegans was included in the apoE4 sequence ("Worm 2"), while in the other construct no signal sequence was added to the apoE4 sequence ("Worm 1").

[0129] [Table 2]

[0130] Extrachromosomal array strains were constructed by injecting the expression plasmid and co-injection marker (pCFJ90, 2 ng / μl for pmyo-2::mCherry) into either Worm 1 or Worm 2. The sequences used for Worm 1 and Worm 2 are included below in Table 2. Coding sequences are shown in uppercase and non-coding sequences are shown in lowercase.

[0131] [Table 3-1] [Table 3-2]

[0132] [Table 4]

[0133] Example 2: An exemplary system for characterizing microbial strains that affect parameters associated with Alzheimer's disease Example 2.1: Alzheimer's Disease Alzheimer's disease (AD) is the most common cause of dementia. AD is characterized by a progressive decline in cognitive functions, including memory, language, and cognitive abilities. Senile plaques and intracellular neurofibrillary tangles are generally considered hallmarks of AD pathology. Plaques can contain aggregates of either 40- or 42-amino acid amyloid-β (Aβ) peptides, which can be formed by abnormal processing of amyloid precursor protein (APP) by presenilins (PSEN1 and PSEN2). Soluble Aβ oligomers can also cause synaptic dysfunction leading to neurodegeneration and cognitive impairment (Mucke, L., and Selkoe, DJ (2012). Neurotoxicity of amyloid β-protein: synaptic and network dysfunction. Cold Spring Harb. Perspect. Med. 2, a006338, which is incorporated herein by reference). Neurofibrillary tangles in AD may contain hyperphosphorylated tau protein, a microtubule-associated protein in neurons (Iqbal, K., et al. (2010). Tau in Alzheimer Disease and Related Tauopathies. Curr. Alzheimer Res. 7, 656-664, which is incorporated herein by reference). In AD, tau may become abnormally hyperphosphorylated and aggregate into fibrils (Grundke-Iqbal, I., et al. (1986). Abnormal phosphorylation of the microtubule-associated protein tau (tau) in Alzheimer cytoskeletal pathology. Proc. Natl. Acad. Sci. USA 83, 4913-4917, which is incorporated herein by reference).Because of the compelling evidence for the involvement of Aβ in AD, several monoclonal antibody-based therapeutics have been developed and tested to target amyloid plaques (van Dyck, CH (2018). Anti-Amyloid-β Monoclonal Antibodies for Alzheimer's Disease: Pitfalls and Promise. Biol. Psychiatry 83, 311-319, which is incorporated by reference in its entirety). However, studies in standard mammalian models have shown that interventions that effectively block or clear Aβ accumulation in animal models do not improve cognition in human clinical trials.

[0134] Approximately 5% of AD cases appear to have a genetic cause, while 95% of cases are sporadic or late-onset AD of unknown etiology. Less than 1% of AD cases are caused by genetic mutations in genes including APP, PSEN1, and PSEN2. Although several other genes have been linked to AD, one of the most prominent genetic risk factors in AD is apoe (Lambert, J.-C., et al. (2009). Genome-wide association study identifies variants at CLU and CR1 associated with Alzheimer's disease. Nat. Genet. 41, 1094-1099; Shen, L., and Jia, J. (2016). An Overview of Genome-Wide Association Studies in Alzheimer's Disease. Neurosci. Bull. 32, 183-190, each of which is incorporated herein by reference). Apolipoprotein E (APOE), a lipid / cholesterol transporter, is encoded by apoE. In humans, there are three major protein variants, designated APOE2, APOE3, and APOE4, which differ from each other at only two amino acid residues (Mahley, RW (2016). Apolipoprotein E: from cardiovascular disease to neurodegenerative disorders. J. Mol. Med. Berl. Ger. 94, 739-746, which is incorporated herein by reference).People who carry apoe polymorphisms, specifically the apoe4 allele, are significantly more likely to develop not only AD, but also early-onset AD, compared with people who carry either the apoe2 or apoe3 alleles (Roses, AD (1996). Apolipoprotein E alleles as risk factors in Alzheimer's disease. Annu. Rev. Med. 47, 387-400; Strittmatter, WJ, and Roses, AD (1996). Apolipoprotein E and Alzheimer's disease. Annu. Rev. Neurosci. 19, 53-77, each of which is incorporated herein by reference). APOE2 is considered a protective form of APOE, while APOE4 is considered a "toxic" form (Strittmatter and Roses, 1996, which is incorporated herein by reference). APOE4 exacerbates AD-associated brain changes, including increased levels of amyloid deposits, brain dysfunction, and neurodegeneration (DiBattista, AM, et al. (2016). Alzheimer's Disease Genetic Risk Factor APOE-ε4 Also Affects Normal Brain Function. Curr. Alzheimer Res. 13, 1200-1207, which is incorporated by reference herein). Despite the importance of APOE4 in AD, the molecular mechanisms by which APOE4 promotes AD pathogenesis remain poorly understood (Kanekiyo, T., et al. (2014). ApoE and Aβ in Alzheimer's disease: accidental encounters or partners? Neuron 81, 740-754, which is incorporated by reference in its entirety). APOE4 is thought to contribute to AD pathogenesis through both loss-of-function and gain-of-function mechanisms (DiBattista, 2016; Zepa, L., et al. (2011).ApoE4-Driven Accumulation of Intraneuronal Oligomerized Aβ42 following Activation of the Amyloid Cascade In Vivo Is Mediated by a Gain of Function. Int. J. Alzheimers Dis. 2011, each of which is incorporated herein by reference.

[0135] Earlier studies suggested that APOE isoforms bind to and help clear Aβ (Kim, J., et al. (2009). The role of apolipoprotein E in Alzheimer's disease. Neuron 63, 287-303, which is incorporated herein by reference). Compared to APOE2 and APOE3, APOE4 was suggested to be less efficient at clearing Aβ (Kim, 2009, which is incorporated herein by reference). However, recent studies suggest that APOE competes with Aβ for uptake via the apoE receptor (Verghese, PB, et al. (2013). APOE influences amyloid-β (Aβ) clearance despite minimal APOE / Aβ association in physiological conditions. Proc. Natl. Acad. Sci. USA110, E1807-1816; Yajima, R., et al. (2015). APOE-isoform-dependent cellular uptake of amyloid-β is mediated by lipoprotein receptor LR11 / SorLA. Biochem. Biophys. Res. Commun. 456, 482-488 (each of which is incorporated herein by reference). While all isoforms were able to compete for binding to the APOE receptor, APOE4-expressing cells were less efficient at clearing Aβ. (Verghese, 2013, which is incorporated herein by reference).

[0136] Although the role of APOE4 in Aβ brain pathology is well documented, the impact of APOE4 in tau pathology has only recently begun to be explored. Using a tauopathy model overexpressing 1N4R human tau containing the P301S mutation, it was shown that ApoE4 exacerbates tau-induced neuroinflammatory and neurodegenerative phenotypes independent of Aβ pathology (Shi, Y., et al. (2017). ApoE4 markedly exacerbates tau-mediated neurodegeneration in a mouse model of tauopathy. Nature 549, 523-527, which is incorporated herein by reference). The tau P301S mutation was originally found in a human case with frontotemporal dementia and degeneration (Bugiani, O., et al. (1999). Frontotemporal Dementia and Corticobasal Degeneration in a Family with a P301S Mutation in Tau. J. Neuropathol. Exp. Neurol. 58, 667-677, which is incorporated herein by reference). Furthermore, the tau P301S mutant protein is a more favorable substrate for phosphorylation compared to wild-type tau (Alonso, A. del C., et al. (2004). Promotion of hyperphosphorylation by frontotemporal dementia tau mutations. J. Biol. Chem. 279, 34873-34881, which is incorporated herein by reference). Interestingly, the neurofibrillary tangles found in AD are primarily composed of hyperphosphorylated tau. (Iqbal, 2010, which is incorporated herein by reference).Also, the frequency of the APOE4 allele is significantly higher in patients with frontotemporal dementia (Stevens, M., et al. (1997). Apolipoprotein E gene and sporadic frontal lobe dementia. Neurology 48, 1526-1529, which is incorporated herein by reference) and APOE4 carriers also have increased disease severity (Agosta, F., et al. (2009). Apolipoprotein E ε4 is associated with disease-specific effects on brain atrophy in Alzheimer's disease and frontotemporal dementia. Proc. Natl. Acad. Sci. 106, 2018-2022; Engelborghs, S., et al. (2006). Dose dependent effect of APOE epsilon4 on behavioral symptoms in frontal lobe dementia. Neurobiol. Aging 27, 285-292, each of which is incorporated herein by reference. Despite the importance of APOE4 in AD, there is a lack of APOE4-targeted therapies (Michaelson, DM (2014). APOE ε4: The most prevalent yet understudied risk factor for Alzheimer's disease. Alzheimers Dement. J. Alzheimers Assoc. 10, 861-868; Holtzman, DM, et al. (2012). Apolipoprotein E and Apolipoprotein E Receptors: Normal Biology and Roles in Alzheimer Disease. Cold Spring Harb. Perspect. Med. 2, each of which is incorporated herein by reference).Moreover, ApoE4 carriers are often excluded from clinical trials for AD due to the unpredictability of their response, despite being identified in more than half of all AD patients (Qiu,WQ,et al.(2013).Angiotensin converting enzyme inhibitors and the reduced risk of Alzheimer's disease in the absence of apolipoprotein E4 allele.J.Alzheimers Dis.JAD 37,421-428, Sperling,R.,et al.(2012).Amyloid-related imaging abnormalities in patients with Alzheimer's disease treated with bapineuzumab:a retrospective analysis.Lancet Neurol.11,241-249, Farlow,MR,et al.(1998).Treatment outcome of tacrine therapy depends on apolipoprotein genotype and gender of the subjects with Alzheimer's disease.Neurology 50,669-677, Risner,ME,et al. (2006). Efficacy of rosiglitazone in a genetically defined population with mild-to-moderate Alzheimer's disease. Pharmacogenomics J. 6, 246-254, each of which is incorporated herein by reference.

[0137] Interestingly, although APOE4 is expressed in the brain, peripheral tissue expression of APOE4 is high, raising the possibility that peripheral APOE4 may contribute to AD pathogenesis. Apart from the brain, APOE protein is mainly synthesized in the liver and is involved in lipid transport and cholesterol homeostasis (Safieh, M., et al. (2019). ApoE4: an emerging therapeutic target for Alzheimer's disease. BMC Med. 17, which is incorporated herein by reference). The liver is the main site that encounters not only nutrients but also small molecules or metabolites or toxins derived from the gut microbiome through the enterohepatic circulation. Thus, gut dysbiosis will have profound effects on the liver. One possibility is that AD may have an intestinal origin, i.e., "microbiome-derived material" may leak into the enterohepatic circulation and reach the liver. From the liver, APOE4 may transport these "microbiome-derived materials" to the brain, where they may seed amyloid deposits and / or increase neuroinflammation. Recent studies have suggested that the gut microbiome plays an important role in AD. Significant changes in the microbiome have been observed in human AD patients compared to control populations. However, it is not known whether these changes are a cause or a consequence of the disease. Although many of the microbiome components have been associated with either susceptibility or pathogenesis of AD, the molecular mechanisms of such interactions remain unknown.

[0138] Example 2.2: APOE4 enhances Aβ-induced paralytic phenotype To test whether microbes modulate AD pathogenesis, transgenic C. elegans strains expressing human APOE4 were grown. These transgenic C. elegans strains expressed human APOE4 in the C. elegans intestine under the control of an intestinal promoter with or without a signal sequence that allows human APOE4 to be secreted from the cell [e.g., mbEx2(pvha-6::ssapoe4)]. In C. elegans, the liver is absent and the intestine performs all the functions that the liver typically performs. Animals were administered standard E. coli OP50 laboratory strain. Animals were monitored every other day from adult day 1 until all animals were paralyzed. For each assay, at least 20 animals (as listed in Table 4) were scored. Data from three independent trials were obtained. For each data point, the mean ± standard deviation is presented in the graph in Figure 1. Transgenic animals expressing human APOE4 with or without the signal sequence did not show any apparent phenotype (Figure 1).

[0139] Expression of human Aβ1-42 in muscle of C. elegans has been reported to induce a paralytic phenotype (Link, CD (1995). Expression of human beta-amyloid peptide in transgenic Caenorhabditis elegans. Proc. Natl. Acad. Sci. USA 92, 9368-9372, which is incorporated herein by reference). To determine whether human APOE4 regulates the paralytic phenotype induced by human Aβ1-42 in muscle, the animals listed in Table 4 were analyzed.

[0140] [Table 5]

[0141] Expression of APOE4 with a signal sequence augmented the dAβ-induced paralytic phenotype. By adult day 8, approximately 40% of animals expressing Aβ were paralyzed, whereas >90% of animals expressing both Aβ and APOE4 with a signal sequence were paralyzed (Figure 1). However, the paralytic phenotype of animals expressing Aβ and APOE4 without a signal sequence was similar to that of animals expressing Aβ alone (Figure 1). In contrast, expression of APOE4 alone in the absence of Aβ expression, with or without a signal sequence, did not induce a paralytic phenotype in adulthood (Figure 1).

[0142] For the remainder of the studies described herein, a line that expressed APOE4 with a signal sequence (which we will refer to as ssApoE4) was analyzed.

[0143] Example 2.3: Aβ3-42 conjugated to GFP and human ssAPOE4 were significantly increased Human Aβ conjugated to GFP in C. elegans muscle 3-42 It has been reported that expression of human ssApoE4 induces Aβ aggregate formation (Link, CD, Fonte, V., Roberts, CM, Hiester, B., Silverman, MA, and Stein, GH (2008). The beta amyloid peptide can act as a modular aggregation domain. Neurobiol. Dis. 32, 420-425, which is incorporated herein by reference). To determine whether expression of human ssApoE4 affects Aβ aggregate formation in C. elegans, human Aβ conjugated to GFP was synthesized. 3-42 The animals listed in Table 5 below were challenged with a standard E. coli OP50 laboratory strain.

[0144] [Table 6]

[0145] When animals reached adulthood, GFP aggregates in the anterior part of the animals were counted. At least 17 animals were recorded for each assay. For each data point, the mean ± standard deviation is presented in the graph in Figure 2. Compared to animals expressing Aβ3-42, animals expressing Aβ3-42 and ssApoE4 had significantly increased GFP aggregates when analyzed using Student's t test (P<0.0001).

[0146] Human Aβ conjugated to GFP 3-42 The number of aggregates in the anterior part of animals expressing Aβ conjugated to GFP compared to that of animals expressing 3-42 The number of aggregates in animals expressing both human ssApoE4 and human ssApoE4 was significantly increased (Figure 2). Interestingly, it has been reported that Aβ deposition in AD patients is higher in APOE4 carriers compared to non-carriers (Dorey, E., Chang, N., Liu, QY, Yang, Z., and Zhang, W. (2014). Apolipoprotein E, amyloid-beta, and neuroinflammation in Alzheimer's disease. Neurosci. Bull. 30, 317-330 (the same reference is incorporated herein by reference)).

[0147] Example 2.4: UbV-GFP is stabilized in animals expressing both human ssAPOE4 and human tau352 (PHP) Hyperphosphorylated tau has been reported to be associated with AD. Expression of pseudohyperphosphorylated tau, which mimics the AD-related modifications, was further reported to induce progressive age-dependent motor impairment in C. elegans (Brandt, R., Gergou, A., Wacker, I., Fath, T., and Hutter, H. (2009). A Caenorhabditis elegans model of tau hyperphosphorylation: induction of developmental defects by transgenic overexpression of Alzheimer's disease-like modified tau. Neurobiol. Aging 30, 22-33, which is incorporated herein by reference).

[0148] To analyze whether ssAPOE4 modulates tau-induced impairment in C. elegans, transgenic strains were generated that expressed mock hyperphosphorylated human tau and human ssAPOE4. Animals of the appropriate genotype were administered standard E. coli OP50 laboratory strains. No apparent differences in locomotion were observed between strains expressing mock hyperphosphorylated human tau and human ssAPOE4 compared to strains expressing mock hyperphosphorylated human tau alone (data not shown).

[0149] Proper proteasome function is important for cellular function, and previous work in the field has shown that proteasome function is impaired in human AD (Bonet-Costa, V., et al. (2016). The Proteasome and Oxidative Stress in Alzheimer's Disease. Antioxid. Redox Signal. 25, 886-901; Upadhya, SC, and Hegde, AN (2007). Role of the ubiquitin proteasome system in Alzheimer's disease. BMC Biochem. 8, S12; Oddo, S. (2008). The ubiquitin-proteasome system in Alzheimer's disease. J. Cell. Mol. Med. 12, 363-373; Zheng, Q., et al. (2016). Dysregulation of Ubiquitin-Proteasome System in Neurodegenerative Diseases. Front. Aging Neurosci. 8, each of which is incorporated herein by reference. To determine whether ssApoE4 expression affects proteasome function, animals carrying human ssApoE4 and markers of impaired proteasome function were generated (Table 6). The proteasome dysfunction marker consisted of non-cleavable ubiquitin fused at the N-terminus to GFP (UbV-GFP).

[0150] [Table 7]

[0151] When the animals reached adulthood, the number of animals expressing GFP in the intestine was counted. At least 30 animals were recorded for each assay. Data from three independent trials are presented in (Figure 3). For each data point, the mean ± standard deviation is presented in the graph. Compared to animals expressing tau352(PHP), animals expressing ssAPOE4 and tau352(PHP) had significantly increased UbV-GFP expression levels when analyzed using a Student's t-test (P<0.0001).

[0152] Generally, UbV-GFP undergoes proteasome-dependent degradation, while impaired protein homeostasis leads to GFP stabilization (see, e.g., FIG. 3). Minimal or absent GFP expression was observed in animals expressing human ssApoE4. However, UbV-GFP was stabilized in animals expressing both human ssApoE4 and human pseudohyperphosphorylated human tau (FIG. 3). Expression of pseudohyperphosphorylated human tau did not induce proteasome stress by itself (FIG. 3). This result suggested that expression of human ssApoE4 and human pseudohyperphosphorylated human tau induced proteasome stress.

[0153] Hyperphosphorylated tau has previously been reported to be resistant to proteasome degradation (Poppek, D., Keck, S., Ermak, G., Jung, T., Stolzing, A., Ullrich, O., Davies, KJA, and Grune, T. (2006). Phosphorylation inhibits turnover of the tau protein by the proteasome: influence of RCAN1 and oxidative stress. Biochem. J. 400, 511-520, which is incorporated herein by reference) and tau phosphorylation has been reported to regulate proteasome activity (Ren, Q.-G., Liao, X.-M., Chen, X.-Q., Liu, G.-P., and Wang, J.-Z. (2007). Effects of tau phosphorylation on proteasome activity. FEBS Lett. 581, 1521-1528; Johnson, GVW (2006). Tau phosphorylation and proteolysis: insights and perspectives. J. Alzheimers Dis. JAD 9, 243-250, which are incorporated herein by reference). In a C. elegans transgenic line, human pseudohyperphosphorylated human tau was expressed in neurons, while human ssApoE4 was expressed under the control of an intestinal promoter with a signal sequence that allows it to be secreted from the cells. Induction of UbV-GFP was observed primarily in the intestine of the animals (not shown). The intestine is a large and prominent tissue in C. elegans, which may mask induction of UbV-GFP in other tissues. However, induction of UbV-GFP in the intestine provided an easy visual screen for possible interventions that modify proteasome function.

[0154] Example 2.5: Microbial strains affect Aβ paralysis Animals of the appropriate genotype were administered either E. coli op50 standard laboratory strain or individual microbiome strains. On adult day 4, the number of paralyzed animals was recorded. Data from three independent trials are presented. For each data point, the mean ± standard deviation is presented on the graph. See Table 1 below for the raw data, including the number of animals analyzed per condition. To facilitate rapid screening of the microbiome for modulators, a novel transgenic C.elegans strain was generated that expresses human ssApoE4, human Aβ1-42, human pseudophosphorylated tau, and UbV-GFP proteasome marker. This transgenic strain can be used not only to identify interventions that suppress paralysis, but also for drugs that improve proteasome function. Furthermore, this model can be used to find parameters or features of biological pathways that affect (increase or decrease) paralysis. The parameters or features can be small molecules, metabolites, nucleic acids, proteins, lipids, or even microbiome components. Approximately 1400 individual microbial strains from the human microbiome were administered to animals carrying human ssApoE4, human Aβ1-42, human pseudophosphorylated tau, and UbV-GFP proteasome marker. The degree of paralysis was observed. A group of microbial populations were found to increase the paralytic phenotype in animals expressing human ssApoE4, human Aβ1-42, human pseudophosphorylated tau, and the UbV-GFP proteasome marker (Table 7, Figure 4). The increase in paralysis was dependent on the presence of ssAPOE4, as administration of many of these bacteria to animals expressing human Aβ1-42, human pseudophosphorylated tau, and the UbV-GFP proteasome marker did not increase the paralytic phenotype (Table 7, Figure 4).

[0155] [Table 8-1] [Table 8-2]

[0156] Example 2.6: Microbial strains modulate Aβ3-42::GFP aggregation Human ssApoE4, human Aβ 1-42 Microbial populations observed to increase paralysis in C. elegans animals expressing tau, human pseudophosphorylated tau, and UbV-GFP proteasome markers included Porphyromonas gingivalis (Table 1). P. gingivalis has been identified in the brains of AD patients and linked to neurotoxic tau and amyloid deposits (Dominy, SS, et al. (2019). Porphyromonas gingivalis in Alzheimer's disease brains: Evidence for disease causation and treatment with small-molecule inhibitors. Sci. Adv. 5, which is incorporated herein by reference). In addition, P. gingivalis has been reported to increase ubiquitin loading, suggesting interference with proteasome function (Dominy et al., 2019, which is incorporated herein by reference). Oral administration of P. gingivalis has previously been shown to be sufficient to induce brain infection and the induction of Aβ deposits (Dominy et al., 2019, incorporated herein by reference).

[0157] Animals were administered either standard E. coli OP50 laboratory strain or individual microbiome strains. GFP aggregates in the anterior part of the animals were counted when the animals reached adulthood. GFP aggregates in three animals were counted for each condition. For each data point, the mean ± standard deviation is presented on the graph.

[0158] Administration of P. gingivalis increased Aβ 3-42 ::GFP aggregates. However, Aβ 3-42 ::GFP aggregates express human ssApoE4 and Aβ 3-42The increase was significantly higher in animals expressing both ssApoE4 and ::GFP together (Figure 5). This result suggested that the ssApoE4 genotype has a detrimental effect on increasing the incidence of AD-related symptoms. Thus, the identification of known microbial populations previously associated with AD confirms the validity of the transgenic human ssApoE4, pseudophosphorylated tau, and Aβ-expressing C. elegans platform assay. The results support that the other microbial populations discovered may be factors influencing AD risk in humans.

[0159] Interestingly, some E. coli isolates expressed increased Aβ along with an increased paralytic phenotype in a ssAPOE4-dependent manner. 3-42 ::GFP aggregates (Table 7, Figure 4, Figure 5). This is interesting, at least because C. elegans is fed the standard non-pathogenic E. coli OP50 strain in the laboratory. Previous studies have shown that gram-negative bacterial molecules, especially from E. coli, are associated with AD neuropathology (Zhan, X., Stamova, B., Jin, L.-W., DeCarli, C., Phinney, B., and Sharp, FR (2016). Gram-negative bacterial molecules associate with Alzheimer disease pathology. Neurology 87, 2324-2332, which is incorporated herein by reference). It is possible that these factors are not expressed or are weakly expressed in the E. coli OP50 strain, or this could suggest strain-specific differences. Eleven strains of E. coli were tested, classified as strains that had either a mild, moderate, or severe effect on paralysis (Table 7, Figure 4) or Aβ 3-42 ::GFP aggregates were phenotyped (Figure 5). These data suggest that strain-specific differences in microbial populations may contribute to the incidence or severity of AD.

[0160] E. fergusonii and E. albertii also showed similar trends to E. coli. Some isolates of E. fergusonii and E. albertii increased the paralysis phenotype, while others had either mild or moderate effects (Table 7). This specific effect on paralysis was also observed in isolates of Klebsiella oxytoca, Klebsiella pnuemoniae, and Alcaligenes faecalis. This may be a general trend for other microbial populations as well, but due to the number of strains analyzed, this feature may have been missed. Thus, among other things, the present disclosure teaches that individual strains of a particular microorganism may have differential effects on biological phenotype(s), including, in particular, disease-related phenotype(s). In some embodiments, the present disclosure provides techniques for identifying and / or characterizing specific strains and / or their components or combinations that may achieve a specific effect on biological phenotype.

[0161] Additionally, microbiome samples from apparently healthy donors were analyzed. It is conceivable that microbiome samples from AD patients may yield better trends in identifying strains that may have adverse effects. However, this C.elegans characterization system can be used to assess the patient's microbiome for increased or decreased presence of microbial strains that are disease-associated or alternatively disease-affecting. While metagenomic sequencing of patient populations can identify the diversity of microbial species present in a particular patient or patient population, these methods cannot identify differences in strain levels in patient samples compared to healthy populations. The system of the present invention fills this gap in identifying differences in strain levels in patient(s) and / or patient population(s) that may be important for some diseases or conditions, including AD. Thus, this platform may provide a potential early diagnostic disease predictor. Thus, among other things, the present disclosure provides techniques for defining, assessing, and / or detecting microbes and / or their components or combinations (i.e., microbial signature(s)) that may be associated with a particular disease state. In some embodiments, such microbial signatures can be detected in a patient sample(s) and can be useful, for example, to diagnose disease conditions, to monitor the impact of particular therapies with respect to such disease conditions, etc.

[0162] Example 2.7: Exemplary Microbial Strains That Affect ATP Production Neuro2A cell line was purchased from ATCC and cultured in EMEM medium supplemented with 10% FBS and 1% L-glutamine. Cells were maintained in a 37°C / 5% CO2 incubator. All experiments were performed using only passages 3-7 cells. Neuro2A cells (5 × 10 per well) were cultured in EMEM medium supplemented with 10% FBS and 1% L-glutamine. Cells were maintained in a 37°C / 5% CO2 incubator. All experiments were performed using only passages 3-7 cells. 4Cells) were plated onto 96-well white-walled plates (Corning) and incubated overnight at 37°C / 5% CO2. Each of the 10 bacteria or a combination of all bacteria (CT10) were grown in the following media: supplemented Clostridium broth, peptone yeast extract glucose broth, MRS broth, and Tryptic Soy broth. Bacteria were grown at 10 in PBS. 8 The cells were resuspended in 0.1 CFU and stored at -80°C. 8 CFU of each microorganism (referred to as sample) was added to six wells. The total bacterial combination (CT10) was added to six wells at 10 9 CFU (i.e., 10 8 CFU of each bacteria). For control wells, PBS without bacteria was added. After 16 h of incubation at 37°C / 5% CO2, 2 μM human amyloid β1 -42 was added to all wells except for the untreated control wells. Cells were incubated for 24 hours at 37°C / 5% CO2. Cells were washed three times with PBS and 0.05 ml of Promega CellTiter-Glo and plates were incubated for 1 hour at room temperature protected from light. Luminescence was measured (representing ATP levels) using a microplate reader (Promega discoverer, Promega Corp). ATP levels were normalized to protein content measured by Bradford protein assay kit (ThermoFisher Scientific). 10 μl of sample was added in duplicate to 150 μl of Bradford reagent in a clear 96-well plate, incubated for 5 minutes in the dark at room temperature and absorbance was measured at 600 nm using a microplate reader (Promega Discoverer, Promega Corp.). Normalized luminescence was calculated by dividing the luminescence value by the OD protein absorbance value. The average of triplicate wells for each condition was calculated and ATP% compared to control was calculated.

[0163] As shown in Figure 6, 2 μM human amyloid beta 1-42Treatment of Neuro2A cells with human amyloid beta ("sham treatment") caused a significant decrease in ATP production. 1-42 In the presence of , treatment of Neuro2A cells with Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp. or Acidaminococcus sp. led to a significant increase in ATP production compared to mock-treated cells, while the combination of all bacteria together (CT-10) led to a further significant increase in ATP production.

[0164] [Table 9-1] [Table 9-2] [Table 9-3] [Table 9-4] [Table 9-5] [Table 9-6] [Table 9-7] [Table 9-8]

[0165] Example 3: An exemplary system for characterizing microbial strains that affect the HIF pathway Example 3.1: HIF Pathway The hypoxia inducible factor (HIF) pathway mediates various metabolic and physiological adaptations to reduced intracellular oxygen levels. Activation of the HIF pathway promotes erythropoiesis and angiogenesis to reduce cellular demand for oxygen. Although the HIF pathway is important in cellular stress responses, constitutive activation of HIF-1 leads to angiogenesis in pathologies such as diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, and glaucoma. Therefore, developing regulators of the HIF pathway is essential for the treatment of intraocular neovascular diseases.

[0166] The HIF-1 pathway consists of the HIF transcription factor and a negative regulator, the prolyl hydroxylase EGLN. EGLN functions as an oxygen sensor, and in the presence of oxygen it hydroxylates the HIF α-subunit (HIFα). Hydroxylation of HIFα leads to binding to, among other factors, the von Hippel-Lindau (VHL) E3 ubiquitin ligase, which promotes HIFα degradation (see Figure 7). Under hypoxic conditions, the HIFα protein is stabilized, which promotes the transcriptional activation of genes required for adaptation to low oxygen levels.

[0167] Example 3.2: Constitutively active HIF-1 leads to egg-laying defects In C. elegans, egl-9 encodes the EGLN homolog. In egl-9 loss-of-function (egl-9 lf) C. elegans mutants, HIF-1 (which is a homolog of HIF1α) protein levels are stabilized. Thus, HIF1 protein is constitutively active, which leads to continuous activity of HIF-1 transcriptional target genes. To identify microorganisms that regulate the HIF-1 pathway, egl-9 loss-of-function (egl-9 lf) C. elegans mutants were analyzed. egl-9 lf mutant C. elegans have constitutively active HIF-1, which leads to them having egg-laying defects. Thus, they become egg-expanded in adulthood.

[0168] Example 3.3: Microbial strains affect HIF-1-induced egg-laying defects HIF-1 modulators were identified by screening individual bacterial strains for their ability to suppress the egg-laying defect of egl-9 lf mutants. Wild-type animals lay 8±2 eggs per hour (n=30), whereas egl-9 lf mutants lay 2±1 eggs / hour (n=30). egl-9 lf C. elegans mutants were administered each of the individual microorganisms and the egg-laying rate of each was measured. In this assay, it was found that Gluconacetobacter spp and Bifidobacterium spp significantly increased the egg-laying rate of egl-9 lf C. elegans mutants to 11±2 eggs (n=25) and 8±2 eggs (n=29), respectively (see Table 9). This example demonstrates that microbial strains such as those in Table 9 can modulate HIF-1 and the HIF-1 pathway and can be used to ameliorate pathologies and diseases associated with alterations in the HIF-1 pathway.

[0169] [Table 10]

[0170] Other embodiments It will be appreciated by those skilled in the art that various changes, modifications, and improvements to the present disclosure will be readily apparent to those skilled in the art. Such changes, modifications, and improvements are intended to be part of this disclosure and are intended to be within the spirit and scope of the present invention. Accordingly, the foregoing description and drawings are by way of example only, and any inventions described in this disclosure are further detailed by the appended claims.

[0171] One of ordinary skill in the art will understand the typical standard deviation or error attributable to values ​​obtained in an assay or other process as described herein. Publications, websites, and other reference materials referenced herein to describe the background of the invention and to provide additional details regarding its practice are hereby incorporated by reference in their entireties. While embodiments of the invention have been described in conjunction with the detailed description, it should be understood that the above description is intended to be illustrative and not limiting of the scope of the invention, which is defined by the appended claims. Other aspects, advantages, and modifications are within the scope of the appended claims. Further aspects of the invention are described below: [Section 1] A system comprising a plurality of C. elegans cultures, each culture comprising a transgenic C. elegans strain that models a mammalian disease or pathology. [Section 2] The plurality of C. elegans cultures (a) 5 or more C. elegans cultures; (b) 10 or more C. elegans cultures; (c) 25 or more C. elegans cultures; or (d) 50 or more C. elegans cultures; 2. The system of claim 1 , comprising: [Section 3] 3. The system of claim 1 or 2, wherein one or more of the cultures comprises a transgenic C. elegans strain that models a human disease or pathology. [Section 4] The system of any one of claims 1 to 3, wherein one or more of the cultures comprises a transgenic C. elegans strain that models Alzheimer's disease. [Section 5] The system of any one of claims 1 to 3, wherein one or more of the cultures comprises a transgenic C. elegans strain that models a disease or pathology associated with an altered or defective HIF-1 pathway. [Section 6] The system of claim 5, wherein the disease or condition associated with an altered or defective HIF-1 pathway comprises a cellular stress response. [Section 7] The system of claim 5, wherein the disease or condition associated with an altered or defective HIF-1 pathway is diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, and glaucoma. [Section 8] The system of claim 5, wherein the disease or condition associated with an altered or defective HIF-1 pathway is an intraocular neovascular disease. [Section 9] 3. The system of claim 1 or 2, wherein one or more of the cultures comprises a transgenic C. elegans strain that models a canine, feline, equine, bovine, ovine, caprine, or porcine disease or condition. [Section 10] 10. The system of claim 1, wherein one or more of the cultures comprises a transgenic C. elegans strain comprising a transgene that includes a reporter gene. [Section 11] The system of claim 10, wherein the reporter gene encodes a fluorescent, phosphorescent, or bioluminescent protein. [Section 12] The system of any one of claims 1 to 11, wherein one or more of the cultures comprises a transgenic C. elegans strain comprising a transgene that includes a mammalian gene associated with a mammalian disease or pathology. [Section 13] The system of any one of claims 1 to 12, wherein one or more of the cultures comprises a transgenic C. elegans strain comprising a transgene comprising a mammalian DNA regulatory element associated with a mammalian disease or pathology. [Section 14] 14. The system of claim 13, wherein the mammalian DNA regulatory element is or comprises an enhancer, promoter, silencer, insulator, locus control region, or a combination thereof. [Section 15] The system of any one of claims 1 to 14, wherein one or more of the cultures comprises a transgenic C. elegans strain comprising a transgene encoding a mammalian RNA regulatory element associated with a mammalian disease or pathology. [Section 16] 16. The system of claim 15, wherein the mammalian RNA regulatory element is or comprises an untranslated region, an intron, a splice site, or a combination thereof. [Section 17] The system of any one of claims 1 to 16, wherein two or more of the cultures comprise transgenic C. elegans strains that model the same mammalian disease or condition. [Section 18] 18. The system of claim 1, wherein all of the cultures comprise transgenic C. elegans strains that model the same mammalian disease or condition. [Section 19] The system of any one of claims 1 to 18, wherein all of the cultures contain the same transgenic C. elegans strain. [Section 20] 18. The system of any one of claims 1 to 17, wherein two or more of the cultures comprise transgenic C. elegans strains that model different mammalian diseases or pathologies. [Section 21] 21. The system of claim 1, wherein one or more of the cultures comprises microorganisms of the mammalian microbiome. [Section 22] 22. The system according to claim 1, wherein each of the cultures comprises microorganisms of the mammalian microbiome. [Section 23] 22. The system of claim 1 , wherein one or more of the cultures comprises microorganisms of the human microbiome. [Section 24] 24. The system of claim 23, wherein each of the cultures comprises microorganisms of the human microbiome. [Section 25] 22. The system of any one of claims 1 to 21, wherein one or more of the cultures comprises microorganisms of the canine, feline, equine, bovine, ovine, caprine, or porcine microbiome. [Section 26] 26. The system of any one of claims 21 to 25, wherein the microbiome is a skin microbiome, an oral microbiome, a nasal microbiome, a gastrointestinal microbiome, a brain microbiome, a lung microbiome, a microbiome, or a urogenital microbiome. [Section 27] 27. The system according to any one of claims 21 to 26, wherein the microorganisms in each culture constitute one or more microbial strains. [Section 28] 28. The system according to any one of claims 21 to 27, wherein the microorganisms in each culture constitute a single microbial strain. [Section 29] 29. The system according to any one of claims 1 to 28, wherein one or more of said cultures constitute a therapeutic or nutritional supplement. [Section 30] A method comprising adding microorganisms obtained from a mammalian microbiome to each of the cultures of the system according to any one of claims 1 to 20. [Section 31] 31. The method of claim 30, wherein the microorganisms added to each culture constitute one or more microbial strains. [Section 32] 32. The method of claim 30 or 31, wherein the microorganisms added to each culture constitute a single microbial strain. [Section 33] 33. The method of any one of claims 30 to 32, wherein the mammalian microbiome is a skin microbiome, an oral microbiome, a nasal microbiome, a gastrointestinal microbiome, a brain microbiome, a lung microbiome, a microbiome, or a urogenital microbiome. [Section 34] 34. The method of any one of claims 30 to 33, wherein one or more of the cultures constitute a therapeutic or nutritional supplement. [Section 35] 35. The method of any one of claims 30 to 34, further comprising determining one or more parameters of the transgenic C. elegans strain in each of the cultures, wherein the one or more parameters are related to the mammalian disease or pathology modeled by the transgenic C. elegans strain. [Section 36] 36. The method of claim 35, wherein the one or more parameters comprise levels of a small molecule, protein, polypeptide, or transcript. [Section 37] The method of claim 35 or 36, wherein the one or more parameters comprise a level of activity of the transgenic C. elegans strain. [Section 38] adding a plurality of microbial strains of a mammalian microbiome to a plurality of C. elegans cultures; A different microbial strain was added to each C. elegans culture. The method, wherein each culture contains the same transgenic C. elegans strain, said transgenic C. elegans strain being a model for a mammalian disease or condition. [Section 39] 39. The method of claim 38, further comprising determining whether each of the plurality of microbial strains affects one or more parameters of the transgenic C. elegans strain, wherein the one or more parameters are associated with the mammalian disease or condition that the transgenic C. elegans strain models. [Section 40] determining one or more parameter values ​​of the transgenic C. elegans strain in a C. elegans culture prior to adding the microbial strain to the culture; determining the same one or more parameter values ​​of the transgenic C. elegans strain in the culture after adding the microbial strain to the C. elegans culture; comparing said one or more parameter values ​​determined before adding said microbial strain with said one or more parameter values ​​determined after adding said microbial strain; 40. The method of claim 38 or 39, further comprising: [Section 41] 41. The method of claim 39 or 40, wherein the one or more parameters comprise levels of a small molecule, a protein, a polypeptide, or a transcript. [Section 42] 42. The method of any one of claims 39 to 41, wherein the one or more parameters comprise a level of activity of the transgenic C. elegans strain. [Section 43] 43. The method of any one of claims 38 to 42, wherein the mammalian microbiome is a skin microbiome, an oral microbiome, a nasal microbiome, a gastrointestinal microbiome, a brain microbiome, a lung microbiome, a microbiome, or a urogenital microbiome. [Section 44] The method of any one of claims 38 to 43, wherein the plurality of C. elegans cultures constitute a therapeutic or nutritional supplement. [Section 45] 45. The method of any one of claims 38 to 44, wherein the transgenic C. elegans strain is a model for a human disease or pathology. [Section 46] 46. ​​The method of any one of claims 38 to 45, wherein the transgenic C. elegans strain is a model for Alzheimer's disease. [Section 47] The method of any one of claims 38 to 45, wherein one or more of the cultures comprises a transgenic C. elegans strain that models a disease or condition associated with an altered or defective HIF-1 pathway. [Section 48] 48. The method of claim 47, wherein the disease or condition associated with an altered or defective HIF-1 pathway comprises a cellular stress response. [Section 49] 48. The method of claim 47, wherein the disease or condition associated with an altered or defective HIF-1 pathway is diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma. [Section 50] 48. The method of claim 47, wherein the disease or condition associated with an altered or defective HIF-1 pathway is an intraocular neovascular disease. [Section 51] 1. A method for characterizing microbial strains of a human biome, comprising: adding the microbial strain to a C. elegans culture, including a transgenic C. elegans strain that models Alzheimer's disease; and determining whether the microbial strain affects one or more parameters of the transgenic C. elegans strain, wherein the one or more parameters are associated with Alzheimer's disease. [Section 52] determining one or more parameter values ​​of the transgenic C. elegans strain in the culture prior to adding the microbial strain to the C. elegans culture; determining the same one or more parameter values ​​of the transgenic C. elegans strain in the culture after adding the microbial strain to the C. elegans culture; comparing said one or more parameter values ​​determined before adding said microbial strain with said one or more parameter values ​​determined after adding said microbial strain; 52. The method of claim 51, further comprising: [Section 53] 53. The method of claim 51 or 52, wherein the transgenic C. elegans strain comprises a transgene encoding human ssApoE4 protein, human Aβ1-42 polypeptide, or human pseudophosphorylated tau protein. [Section 54] The one or more parameters are (i) the level of C. elegans paralysis; (ii) amyloid plaque levels; (iii) levels of tau fibrils; (iv) the level of neuroinflammation; (v) the level of proteasome function, or (vi) any combination thereof; The method according to any one of claims 51 to 53, comprising: [Section 55] 1. A method for characterizing microbial strains of a human biome, comprising: adding the microbial strain to a C. elegans culture, including a transgenic C. elegans strain that models a disease or condition associated with an altered or defective HIF-1 pathway; and determining whether the microbial strain affects one or more parameters of the transgenic C. elegans strain, wherein the one or more parameters are associated with an altered or defective HIF-1 pathway. [Section 56] determining one or more parameter values ​​of the transgenic C. elegans strain in the culture prior to adding the microbial strain to the C. elegans culture; determining the same one or more parameter values ​​of the transgenic C. elegans strain in the culture after adding the microbial strain to the C. elegans culture; comparing said one or more parameter values ​​determined before adding said microbial strain with said one or more parameter values ​​determined after adding said microbial strain; 56. The method of claim 55, further comprising: [Section 57] 57. The method of claim 55 or 56, wherein the transgenic C. elegans strain comprises a transgene encoding the human prolyl hydroxylase EGLN, a human HIF transcription factor, or a human HIFα protein. [Section 58] The one or more parameters are (i) the level of neuroinflammation; (ii) the level of proteasome function; (iii) the level of C. elegans egg laying rate, or (iv) combinations thereof; The method according to any one of claims 55 to 57, comprising: [Section 59] 21. Use of a system according to any one of claims 1 to 20 for screening a mammalian microbiome for microbial strains that influence a mammalian disease or pathology. [Section 60] 60. The use of claim 59, wherein the mammalian microbiome is a human microbiome. [Section 61] 21. Use of a system according to any one of claims 1 to 20 for characterising the impact that microbial strains of a mammalian microbiome have on a mammalian disease or pathology. [Section 62] Transgenic C. elegans lines expressing two or more of: (i) human ssApoE4, (ii) human Aβ1-42, (iii) human pseudophosphorylated tau, and (iv) UbV-GFP proteasome marker. [Section 63] A composition comprising one or more microbial strains listed in Table 8. [Section 64] 64. The composition of claim 63, comprising two or more microbial strains listed in Table 8. [Section 65] 64. The composition of claim 63, comprising five or more microbial strains listed in Table 8. [Section 66] 64. The composition of claim 63, comprising 10 or more microbial strains listed in Table 8. [Section 67] A composition comprising Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium sp., Bacillus subtilis, Acidaminococcus sp., or combinations thereof. [Section 68] 68. The composition of claim 67, comprising at least two microbial strains selected from the group consisting of Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium, Bacillus subtilis, and Acidaminococcus sp. [Section 69] 68. The composition of claim 67, comprising at least five microbial strains selected from the group consisting of Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium, Bacillus subtilis, and Acidaminococcus sp. [Section 70] 68. The composition of claim 67, comprising Gluconacetobacter hansenii, Terrisporobacter glycolicus, Coprococcus sp., L. plantarum, Clostridium butyricum, Paenibacillus sp., Veillonella sp., Bifidobacterium, Bacillus subtilis, and Acidaminococcus sp. [Section 71] The composition according to any one of claims 63 to 70, wherein the composition is a pharmaceutical composition. [Section 72] 71. The composition of any one of claims 63 to 70, wherein the composition is an ingestible item. [Section 73] 73. A method of treating a disease or condition in a subject, comprising administering to a subject in need thereof a composition according to any one of claims 63 to 72. [Section 74] 74. The method of claim 73, wherein the disease or condition is a neurodegenerative disease or disorder. [Section 75] 74. The method of claim 73, wherein the disease or condition is Alzheimer's disease. [Section 76] 74. The method of claim 73, wherein the disease or condition is associated with an altered or defective HIF-1 pathway. [Section 77] 74. The method of claim 73, wherein the disease or condition is an intraocular neovascular disease or disorder. [Section 78] 74. The method of claim 73, wherein the disease or condition is diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma. [Section 79] 73. Use of a composition according to any one of claims 63 to 72 in the treatment of a disease or condition in a subject. [Section 80] 80. The use of claim 79, wherein the disease or condition is a neurodegenerative disease or disorder. [Section 81] 80. The use of claim 79, wherein the disease or condition is Alzheimer's disease. [Section 82] 80. The use of claim 79, wherein the disease or condition is associated with an altered or defective HIF-1 pathway. [Section 83] 80. The use of claim 79, wherein the disease or condition is an intraocular neovascular disease or disorder. [Section 84] 80. The use of claim 79, wherein the disease or condition is diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, or glaucoma.

Claims

1. A system for evaluating one or more microorganisms of the human microbiome, comprising: (i) a plurality of C. elegans cultures; wherein one or more of said cultures comprises a transgenic C. elegans strain that models a mammalian disease or condition; the mammalian disease or condition is Alzheimer's disease; The transgenic C. elegans strain contains the human ApoE4 gene, human Aβ １－４２ Gene, human Aβ ３－４２ a transgene comprising a gene, a human tau gene, a human pseudophosphorylated tau gene, or a pseudohyperphosphorylated human tau gene, and (ii) one or more microorganisms of the human microbiome to be evaluated wherein each of said cultures comprises one or more microorganisms of the human microbiome.

2. The plurality of C. elegans cultures (a) 5 or more C. elegans cultures; (b) 10 or more C. elegans cultures; (c) 25 or more C. elegans cultures; or (d) 50 or more C. elegans cultures; 2. The system of claim 1 , comprising:

3. 3. The system of claim 1 or 2, wherein one or more of the cultures comprises a transgenic C. elegans strain that contains a transgene that includes a reporter gene.

4. 4. The system of any one of claims 1 to 3, wherein one or more of the cultures comprises a transgenic C. elegans strain comprising a transgene comprising a mammalian DNA regulatory element associated with a mammalian disease or pathology.

5. 4. The system of any one of claims 1 to 3, wherein one or more of the cultures comprises a transgenic C. elegans strain comprising a transgene encoding a mammalian RNA regulatory element associated with a mammalian disease or condition.

6. 6. The system of any one of claims 1 to 5, wherein two or more of the cultures comprise transgenic C. elegans strains that model the same mammalian disease or condition.

7. 7. The system of any one of claims 1 to 6, wherein all of the cultures comprise transgenic C. elegans strains that model the same mammalian disease or condition.

8. 8. The system of any one of claims 1 to 7, wherein all of the cultures contain the same transgenic C. elegans strain.

9. 6. The system of any one of claims 1 to 5, wherein two or more of the cultures comprise transgenic C. elegans strains that model different mammalian diseases or pathologies.

10. 10. The system of any one of claims 1 to 9, wherein the microbiome is a skin microbiome, an oral microbiome, a nasal microbiome, a gastrointestinal microbiome, a brain microbiome, a lung microbiome, or a urogenital microbiome.

11. 11. The system according to any one of claims 1 to 10, wherein the microorganisms in each culture comprise one or more microbial strains.

12. 11. The system according to any one of claims 1 to 10, wherein the microorganisms in each culture comprise a single strain of microorganism.

13. The system of any one of claims 1 to 12, wherein one or more of the cultures comprises a therapeutic or nutritional supplement.

14. A method for the preparation of a culture medium comprising adding one or more microorganisms obtained from a human microbiome to each of the cultures in a system, the system comprises a plurality of C. elegans cultures; one or more of said cultures comprises a transgenic C. elegans strain that models a mammalian disease; the mammalian disease is Alzheimer's disease, and The transgenic C. elegans strain contains the human ApoE4 gene, human Aβ １－４２ Gene, human Aβ ３－４２ The method includes a transgene comprising a gene, a human tau gene, a human pseudophosphorylated tau gene, or a pseudohyperphosphorylated human tau gene.

15. 15. The method of claim 14, wherein the microorganisms added to each culture comprise one or more microbial strains.

16. 15. The method of claim 14, wherein the microorganisms added to each culture comprise a single microbial strain.

17. 17. The method of any one of claims 14 to 16, wherein one or more of the cultures comprises a therapeutic or nutritional supplement.

18. 18. The method of any one of claims 14 to 17, further comprising determining one or more parameters of the transgenic C. elegans strain in each of the cultures, the one or more parameters being associated with the mammalian disease or condition that the transgenic C. elegans strain models, and comprising a protein amount, a polypeptide amount, or a transcript amount.

19. 18. The method of any one of claims 14 to 17, further comprising determining one or more parameters of the transgenic C. elegans strain in each of the cultures, the one or more parameters being associated with the mammalian disease or condition that the transgenic C. elegans strain models, and comprising a level of activity of the transgenic C. elegans strain.

20. A method for the preparation of a culture of C. elegans comprising adding a plurality of microbial strains of the human microbiome to a plurality of C. elegans cultures, A different microbial strain was added to each C. elegans culture, Each culture contained the same transgenic C. elegans strain, The transgenic C. elegans strain serves as a model for Alzheimer's disease; and The transgenic C. elegans strain contains the human ApoE4 gene, human Aβ １－４２ Gene, human Aβ ３－４２ The method includes a transgene comprising a gene, a human tau gene, a human pseudophosphorylated tau gene, or a pseudohyperphosphorylated human tau gene.

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