WO2018075886A1

WO2018075886A1 - Compositions and methods for inducing immune system maintenance to prevent and/or treat infections

Info

Publication number: WO2018075886A1
Application number: PCT/US2017/057581
Authority: WO
Inventors: Jack A. GILBERT; John C. Alverdy; Daniel Van Der Lelie; Safiyh Taghavi
Original assignee: The University Of Chicago; Gusto Global, Llc
Priority date: 2016-10-21
Filing date: 2017-10-20
Publication date: 2018-04-26

Abstract

Provided herein are compositions (e.g., probiotic, therapeutics, pharmaceutical, etc.) comprising one or more strains of bacteria from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae. In particular, provided herein are compositions comprising bacteria from one or more of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus and methods of use thereof for inducing immune system maintenance and/or rescuing animals from sepsis.

Description

COMPOSITIONS AND METHODS FOR INDUCING IMMUNE SYSTEM MAINTENANCE TO PREVENT AND/OR TREAT INFECTIONS

CROSS REFERENCE TO RELATED APPLICATIONS

The present application claims the priority benefit of U. S. Provisional Patent

Application No. 62/410,897, filed October 21, 2016, which is incorporated by reference in its entirety.

STATEMENT REGARDING FEDERAL FUNDING

This invention was made with government support under R01 GM062344 awarded by

The National Institute of Health (NIH). The government has certain rights in the invention

FIELD

Provided herein are compositions (e.g., probiotic, therapeutics, pharmaceutical, etc.) comprising one or more strains of bacteria from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae and/or Bacteroidaceae. In particular embodiments, provided herein are compositions comprising bacteria from one or more of the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, and methods of use thereof for inducing immune system maintenance for preventing and/or rescuing humans and animals from infections, including sepsis.

BACKGROUND

Exposure to the physiologic stress of critical illness and injury has a profound effect on the intestinal microbiome, physicochemical characteristics of the gut microenvironment, and local barrier function. These effects can be induced by both the host stress response and the selective pressures of modem intensive care therapy, and one potential consequence is the loss of colonization resistance to highly virulent and antibiotic-resistant healthcare acquired pathogens that can rapidly colonize the gut (Kanazawa 2005, Shimizu 2006, Shimizu 2011 , Zaborin et al, MBio 2014; incorporated by reference in their entireties). When the local immune system is not able to contain these pathogens, dissemination into the bloodstream and peripheral organs occurs, leading to a systemic immune response that on one hand, may drive the clearance of pathogens, but when inappropriately sustained, can cause organ dysfunction and ultimately lead to mortality. Despite continuing efforts to develop treatments and clarify its underlying biology, sepsis remains the most common cause of death in the modern intensive care unit, and given the persisting mortality rates and reportedly increasing prevalence (Angus et al, Crit Care Med 2001 ; Lagu et al, Crit Care Med 2012; incorporated by reference in their entireties), the discovery and validation of novel therapeutic agents is imperative.

First chronicled in 4th century China, the use of feces from healthy donors to treat disease, or fecal microbiota transplantation (FMT), is a therapy that has had several historical iterations across multiple medical disciplines (Zhang et al., Am J Gastroenterol 2012;

incorporated by reference in its entirety). In modern medicine, FMT resurfaced as a potential treatment for pseudomembranous colitis (Eiseman et al., Surgery 1958; incorporated by reference in its entirety), and has recently been tested for therapeutic effects against inflammatory bowel disease (Colman and Rubin, J Crohns Colitis. 2014; Pigneur and Sokol, Mucosal Immunol 2016; incorporated by reference in their entireties), metabolic syndrome (Vrieze et al, Gastroenterology 2012; incorporated by reference in its entirety), and other disorders (Smits et al., Gastroenterology 2013; incorporated by reference in its entirety) to varying degrees of efficacy. Notably, recurrent Clostridium difficile infection is accompanied by a severe reduction in gut microbial diversity, and multiple studies have shown that treatment of patients with C. difficile using FMT leads to a restoration of gut microbial diversity accompanied by a significant increase in cure rate (van Nood et al., NEJM 2013; Cammarota et al, J Clin Gastroenterol 2014; incorporated by reference in its entirety).

SUMMARY

Provided herein are compositions (e.g., probiotic, therapeutics, pharmaceutical, etc.) comprising one or more strains of bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae. In particular embodiments, provided herein are compositions comprising bacteria from one or more of the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus and methods of use thereof for inducing immune system maintenance for preventing and/or rescuing humans and animals from infections including sepsis.

In some embodiments, provided herein are methods of modulating an immune response in a subject, the method comprising administering to the subject a composition comprising bacteria from one or more of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus (e.g., selected from the strains listed in Tables 1-6). In some embodiments, the modulation of the immune response comprises (or results in) preventing (prophylactically) and/or treating (responsively) infections, including sepsis. In some embodiments, the subject has an infection that may result in sepsis. In some embodiments, the subject is at risk (e.g., increased risk) of developing an infection that can result in sepsis. In some embodiments, the modulation of the immune response comprises (or results in) treating sepsis. In some embodiments, the subject suffers from sepsis. In some embodiments, the subject suffers from severe sepsis or septic shock. In some embodiments, the composition comprises bacteria from 2 or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more, or ranges therebetween) different taxa (e.g., families, genera, species, strains etc.). In some embodiments, the composition comprises at least 10⁴ colony forming units (CFU) of bacteria (e.g., at least l lO⁴ CFU, 2xl0⁴ CFU, 5x l0⁴ CFU, l x lO⁵ CFU, 2xl0⁵ CFU, 5xl0⁵ CFU, l x lO⁶ CFU, 2xl0⁶ CFU, 5x l0⁶ CFU, l xlO⁷ CFU, 2x l0⁷ CFU, 5xl0⁷ CFU, l xlO⁸ CFU, 2x l0⁸ CFU, 5xl0⁸ CFU, l x lO⁹ CFU, 2xl0⁹ CFU,

5x l0⁹ CFU, l xlO¹⁰ CFU, 2xl0¹⁰ CFU, 5x l0¹⁰ CFU, l x lO¹¹ CFU, 2xlOⁿ CFU, 5xlOⁿ CFU, l x lO¹² CFU, 2x l0¹² CFU, 5xl0¹² CFU, or more or ranges there between). In some embodiments, the subject has abnormal gut microbiota. In some embodiments, the subject is a human. In some embodiments, the subject is an animal (e.g., livestock, domestic pet, research subject, etc.). In some embodiments, the composition is administered orally, topically, rectally, etc. In some embodiments, the composition is co-administered with one or more additional active agents. In some embodiments, the additional active agents are part of the same formulation. In some embodiments, the additional active agents are administered separately as part of the treatment plan for the prevention and/or control of an infection. In some embodiments, the additional active agent comprises a probiotic component or a prebiotic component. In some embodiments, the additional active agent comprises a small molecule based therapeutic. In some embodiments, the additional small molecule is an antibiotic, a signaling compound, or a compound that attenuates the virulence phenotype of pathogenic microbes. In some embodiments, the additional active agent comprises an antibody or antibody fragment. In some embodiments, the additional active agent comprises a peptide or polypeptide. In some embodiments, the additional active agent comprises a microbe.

In some embodiments, methods described herein comprise assaying the microbiome and/or metabolome of a subject. In some embodiments, assaying the microbiome comprises testing the presence, absence, relative abundance or amount of one or more bacteria in the gut of the subject. In some embodiments, assaying the microbiome comprises testing the presence, absence, relative abundance or amount of one or more bacteria (e.g., selected from the strains listed in Tables 1-6) from one or more of the families Prevotellaceae,

Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus. In some embodiments, assaying the metabolome comprises quantifying amount of one or more metabolites in the gut of the subject. In some embodiments, the assaying is performed on the subject before and/or after administration of the composition.

In some embodiments, provided herein are pharmaceutical compositions comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from one or more of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus. In some embodiments, pharmaceutical compositions comprise a therapeutically effective amount of bacteria. In some embodiments, a therapeutically effective amount of bacteria is an amount sufficient to treat or prevent an infection, including sepsis, in a subject. In some

embodiments, a therapeutically effective amount of bacteria is an amount sufficient to activate regulator T cell accumulation in the subject. In some embodiments, the composition comprises at least 10⁴ colony forming units (CFU) of bacteria (e.g., at least 1 * 10⁴ CFU, 2x l0⁴ CFU, 5 x l0⁴ CFU, l x lO⁵ CFU, 2x l0⁵ CFU, 5 x l0⁵ CFU, l x lO⁶ CFU, 2x l0⁶ CFU, 5x l0⁶ CFU, l x lO⁷ CFU, 2x l0⁷ CFU, 5x l0⁷ CFU, l x lO⁸ CFU, 2x l0⁸ CFU, 5 x l0⁸ CFU, l x lO⁹ CFU, 2x l0⁹ CFU, 5 x l0⁹ CFU, l x lO¹⁰ CFU, 2x l0¹⁰ CFU, 5x l0¹⁰ CFU, l x lO¹¹ CFU, 2x lOⁿ CFU, 5x l0ⁿ CFU, l x lO¹² CFU, 2x l0¹² CFU, 5x l0¹² CFU, or more or ranges there between). In some embodiments, pharmaceutical compositions comprise a probiotic or a prebiotic. In some embodiments, the bacteria are alive as vegetative cells and/or spores. In some embodiments, pharmaceutical compositions are formulated for oral, rectal, and/or topical administration. In some embodiments, the pharmaceutical composition is a nutraceutical or a food.

In some embodiments, provided herein is the use of a pharmaceutical composition described herein to manufacture a medicament for administration to a subject (e.g., for the treatment or prevention of an infection, including sepsis). In some embodiments, provided herein is the use of a pharmaceutical composition described herein to treat or prevent an infection, including sepsis, in a subject.

Methods are provided for the treatment of subjects in need of treatment for immune response modulation (e.g., subject with bacterial infections, subjects with abnormal microbiota, subjects suffering from sepsis, etc.). In other embodiments, methods are provided (e.g., prophylactically) for the prevention of development of an infection that could lead to sepsis in a subject (e.g., a subject at increased risk of sepsis). In other embodiments, methods are provided (e.g., prophylactically) for the prevention of development of sepsis from an infection (e.g., in a subject at increased risk of sepsis should they develop an infection). In some embodiments, bacterial compositions described herein (e.g., comprising bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus) are administered to a subject having gut microbiota that places the subject at risk of developing an infection that could lead to sepsis. In some embodiments, bacterial compositions described herein (e.g., comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes,

Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus) are administered to a subject having gut microbiota that has caused the subject to experience inflammatory conditions (e.g., that place the subject at risk for sepsis). In some exemplary embodiments, methods comprise treating a subject who has a gut microbiota that differs from the subject's normal microbiota or microbiota of a healthy individual in one or both of membership or relative abundance of one or more members of the gut microbiota, e.g., methods comprise administering a composition comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae,

Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus to a subject who has a gut microbiota that differs from the subject's normal microbiota or microbiota of a healthy individual, in one or both of membership or relative abundance of one or more members of gut microbiota that are preventative of infections that could result in sepsis. In some embodiments, the invention relates to methods comprising treating a subject that has a gut microbiota that differs from the normal microbiota (e.g., microbiota that promotes healthy immune response) in the membership or relative abundance of bacteria from the families Prevotellaceae, Rikenellaceae,

Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or

Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, e.g., methods comprising

administering a composition comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae,

Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus to a subject who has a gut microbiota that differs from the normal microbiota (e.g., microbiota that promotes healthy immune response (e.g., to a bacterial infection)) in the membership or relative abundance of the beneficial bacteria.

In some embodiments, provided herein are methods of preventing unrestrained immune/inflammatory responses e.g. caused by infection (e.g., a bacterial infection) in a subject, the methods comprising administering (e.g., prophylactically) a composition comprising bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus to the subject. In some embodiments, the subject is at risk (e.g., increased risk) of developing sepsis from an infection (or potential infection), e.g. caused by genetic factors (e.g., determined from genetic testing), abnormal gut microbiota, age, an underlying condition (being immunocompromised, etc.), surgical complications, open wounds.

The technology is not limited in the types or classes of subj ects or patients that are treated and/or that are administered with the compositions herein. For example, in some embodiments the subj ect is a human. In some embodiments, the subject is a young human, e.g., that has not developed a robust or complete gut microbiota that helps to train the immune system, helps to control inflammatory conditions, and/or prevents sepsis. For example, in some embodiments the subject is a human infant or a human neonate or a human newborn. In some embodiments, the subj ect is a juvenile, adolescent, adult, or elderly subject. In some embodiments, the subject has experienced a shock to their microbiota (e.g., antibiotic resistant infection, antibiotic use, organ transplant, chemotherapy, etc.). The technology is applicable to subjects and patients that are nonhuman, e.g., mammals, birds, etc., including but not limited to livestock animals, domesticated animals, animals in captivity, etc. The technology is not limited in the type or route if administration. In some embodiments, the type or route of administration provides the composition comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Rununococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus to the subject's gastrointestinal tract. For example, in some embodiments the composition is administered orally and in some embodiments the composition is administered rectally. In some embodiments, the composition is administered, via inhalation, topically, etc.

Some embodiments comprise administering compositions comprising class bacteria

(e.g., bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families

Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Rununococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, and taxonomically -related bacteria that similarly support healthy immune system and/or treat/prevent infections that can lead to sepsis) and one or more additional components. In some embodiments, additional components are selected from other bacterial and nonbacterial components (e.g., for treatment/prevention of infections that could result in sepsis, for formulation of the composition (e.g., stability, shelf-life, consistency, taste, strain engrafting, strain activity, etc.), etc.).

In some embodiments, the technology comprises testing a subject or a patient. For example, some embodiments comprise testing the subject for the presence, absence, amount or relative abundance of bacteria (e.g., bacteria from the families Prevotellaceae,

Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Rununococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes,

Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus bacteria (e.g., selected from the strains listed in Tables 1 -6)) and/or in the gut microbiota. In some embodiments, a subject is tested for the presence of metabolites that promote healthy immune system, capacity to respond to sepsis or a bacterial infection, etc. Some embodiments comprise testing the subject for an abnormal gut microbiota. The technology provides methods in which a subject or a patient is tested before and/or after administration of a composition comprising bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae Rununococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus to the subject or patient. In some embodiments, the testing informs the dose amount, dose schedule, and/or CFU of bacteria in the composition that is administered to the subject or patient. Some embodiments comprise administration of a composition comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Bacteroides, Lactobacillus,

Parabacteroides, and/or Ruminococcus to the subject or patient, testing the subject or patient, and a second administration of a composition comprising bacteria from the families

Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus to the subject. The first and second administrations and/or compositions may be the same or different, e.g., same or different in dose, amount, route, composition, species of bacteria, CFU of bacteria, type of small therapeutic molecules, etc.

Some embodiments herein include testing the subject or patient for infection (e.g., bacterial, viral, etc.), sepsis, severe sepsis, septic shock, etc.

Some embodiments herein include testing the subject or patient for normal gut microbiota (e.g., microbiota that promotes healthy immune response); abnormal gut microbiota (e.g., microbiota that prevents healthy immune response, pathogenic microbiota, etc.); or presence, absence, number, or relative abundance of specific taxa or strains of bacteria (e.g., bacteria from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, etc.) in the gut microbiota. In some embodiments, such testing comprises: analysis of a biomarker such as a metabolite, nucleic acid, polypeptide, sugar, lipid, indication, symptom, etc. For example, in some embodiments the technology comprises testing using a labeled probe, a nucleic acid test (NAT), a nucleic acid amplification test (NAAT), a nucleic acid amplification technology (e.g., polymerase chain reaction (e.g., PCR, real-time PCR, probe hydrolysis PCR, reverse transcription PCR), isothermal amplification (e.g., nucleic acid sequence-based amplification (NASBA)), a ligase chain reaction, or a transcription mediated amplification, etc.), or nucleic acid sequencing (e.g., Sanger sequencing or next-gen (e.g., second generation, third generation, etc.) sequencing methods including, e.g., sequencing-by-synthesis, single molecule sequencing, nanopore, ion torrent, etc.).

Some embodiments comprise a second testing of the subject or patient (e.g., for microbiota composition, metabolite composition, infection, etc.), which may be the same or different from the first testing of the patient. In some embodiments, the second testing occurs after administration of a composition comprising bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, and/or taxonomically-related bacteria that similarly modulate the immune response and/or treat/prevent infections that could lead to sepsis in the subject. In some embodiments, the second testing indicates that the administration of the composition comprising bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus is an effective treatment. In some embodiments, the second testing indicates that the administration of the composition comprising bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae,

Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera

Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus was an ineffective treatment. In some embodiments, the dose amount, dose schedule, and/or type or CFU of bacteria in the composition is changed for subsequent administrations to the subject or patient based on the results of the test.

In some embodiments, methods comprise administering (e.g., therapeutically or prophylactically) to a subject or patient a composition comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, and a probiotic component, a prebiotic component and/or a small molecule therapeutic compound.

Non-limiting examples of prebiotics useful in the compositions and methods herein include xylose, arabinose, ribose, galactose, rhamnose, cellobiose, fructose, lactose, salicin, sucrose, glucose, esculin, tween 80, trehalose, maltose, mannose, mellibiose, raffinose, fructooligosaccharides (e.g., oligofructose, inulin, inulin-t pe fructans),

galactooligosaccharides, amino acids, alcohols, water-soluble cellulose derivatives (most preferably, methylcellulose, methyl ethyl cellulose, hydroxyethyl cellulose, ethyl

hydroxyethyl cellulose, cationic hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxyethyl methylcellulose, hydroxypropyl methylcellulose, and carboxymethyl cellulose), water-insoluble cellulose derivatives (most preferably, ethyl cellulose), unprocessed oatmeal, metamucil, all-bran, and any combinations thereof.

In some embodiments, methods comprise administering to a subj ect or patient a composition comprising bacteria (e.g., selected from the strains listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae,

Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera

Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, and small molecule therapeutics. Non-limiting examples of small molecule- based therapeutics useful in the compositions and methods herein include an antibiotic, a signaling compound (e.g. to disrupt biofilm formation of pathogens), or a compound that attenuates the virulence phenotype of pathogenic microbes, such as PiPEG15-20 and similar molecules (Zaborin et al. 2014. Antimicrob Agents Chemother. 58: 966-977; incorporated by reference in its entirety).

Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera

Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, and one or more antibody and/or antibody fragment therapeutics. Exemplary antibodies that find use in the treatment of infection include, for example, those described in U. S. Pat. No. 7,569,677; U. S. Pub No. 2010/0272736; incorporated by reference in their entireties.

In some embodiments, the additional active agents are part of the same formulation. In some embodiments when administered as separate formulations, the co-administration of two or more agents or therapies is concurrent. In other embodiments, a first agent/therapy is administered prior to a second agent/therapy as part of the treatment plan for the prevention and/or control of an infection.

Embodiments provide that pharmaceutical compositions are formulated for administration to a subject or a patient, e.g., some embodiments provide pharmaceutical compositions formulated for administration to a human or an animal. Some embodiments provide pharmaceutical compositions formulated for human administration to a human newborn, neonate, infant, juvenile, teen, adult, or elderly patient. Related embodiments provide a pharmaceutical composition comprising live bacteria as vegetative cells and/or bacterial spores.

Embodiments provide pharmaceutical compositions formulated for various routes of administration, e.g., to the gastrointestinal tract. For example, in some embodiments, the pharmaceutical composition is formulated for oral administration and in some embodiments the pharmaceutical composition is formulated for rectal administration. In some embodiments, the pharmaceutical composition is a nutraceutical or a food. In some embodiments, other routes of administration are used (e.g., topical, etc.)

Related embodiments provide kits comprising a pharmaceutical composition comprising bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or from the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus. Some embodiments provide a kit for treating or preventing infections that could lead to sepsis in a subject, the kit comprising a composition comprising bacteria formulated for administration to the subj ect; and a reagent for testing the membership or relative abundance of one or more members of the gut microbiota of the subject. In some embodiments, the kit reagent comprises a labeled oligonucleotide probe. In some embodiments, the kit reagent comprises an amplification oligonucleotide. Embodiments of kits comprise a reagent that provides a test for the presence, absence, or level of specific strains or taxa described herein in the gut microbiota of the subject, etc.

BRIEF DESCRIPTION OF THE DRAWINGS

Figures 1 A-B. Fecal microbial transplant (FMT) rescues mice from mortality in gut- derived polymicrobial sepsis. (Fig. 1A), sketch and timeline of the experimental design demonstrating hepatectomy and cecal inoculation by a pathogen community. (Fig. IB), Kaplan-Meier survival curves, n = 15/group, pO.001 between FMTii_ve vs FMT_aut0d-

Figures 2A-F. Fecal microbial transplant prevents the formation of a pathobiome characterized by a low diversity, pathogen-dominating structure, and pro-inflammatory function. (Fig. 2A), Shannon diversity index. (Fig. 2B), Relative abundance of taxa at genus level. Black columns correspond to Serratia and red columns- to Enterococcus . (Figs. 2C-E), Oligotyping analysis of Serratia marcescens (Fig. 2C), Enterococcus faecalis (Fig. 2D), and Klebsiella oxytoca (Fig. 2E). Black color columns representing oligotype 1 for each of these species are 100 % similarity to oligotypes of community strains derived based on the whole genome sequence data. (Fig. 2F), Cytokine production by splenic dendritic cells in the response to cecal contents of untreated, POD2- FMT_aut0d and POD2- FMTii_ve groups measured by ELISA. n=4-8 mouse cecal contents per group, *p<0.01.

Figures 3 A- J. Effect of FMT on systemic sepsis in IP sepsis model. (Fig. 3 A), Morbidity score. n=10/group, *p<0.05. (Fig. 3B) Kaplan-Meyer survival curves. (Figs. 3C- F), Dissemination at POD2. n=5/group, p<0.05. (Figs. 3G-J), QRT-PCR of RIG-I related genes determined in cecum, liver, and spleen at POD2 in IP model. n=3/group, *p<0.05. For comparison, microarray data of the genes expression in cecum, liver, and spleen in the hepatectomy model are displayed.

DEFINITIONS Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments described herein, some preferred methods, compositions, devices, and materials are described herein. However, before the present materials and methods are described, it is to be understood that this invention is not limited to the particular molecules, compositions, methodologies or protocols herein described, as these may vary in accordance with routine experimentation and optimization. It is also to be understood that the terminology used in the description is for the purpose of describing the particular versions or embodiments only, and is not intended to limit the scope of the embodiments described herein.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. However, in case of conflict, the present specification, including definitions, will control. Accordingly, in the context of the embodiments described herein, the following definitions apply.

As used herein and in the appended claims, the singular forms "a", "an" and "the" include plural reference unless the context clearly dictates otherwise. Thus, for example, reference to "a probiotic agent" is a reference to one or more probiotic agents and equivalents thereof known to those skilled in the art, and so forth. As used herein, the term "and/or" includes any and all combinations of listed items, including any of the listed items individually. For example, "A, B, and/or C" encompasses A, B, C, AB, AC, BC, and ABC, each of which is to be considered separately described by the statement "A, B, and/or C." As used herein, the term "comprise" and linguistic variations thereof denote the presence of recited feature(s), element(s), method step(s), etc. without the exclusion of the presence of additional feature(s), element(s), method step(s), etc. Conversely, the term "consisting of and linguistic variations thereof, denotes the presence of recited feature(s), element(s), method step(s), etc. and excludes any unrecited feature(s), element(s), method step(s), etc., except for ordinarily-associated impurities. The phrase "consisting essentially of denotes the recited feature(s), element(s), method step(s), etc. and any additional feature(s), element(s), method step(s), etc. that do not materially affect the basic nature of the composition, system, or method. Many embodiments herein are described using open "comprising" language. Such embodiments encompass multiple closed "consisting of and/or "consisting essentially of embodiments, which may alternatively be claimed or described using such language.

As used herein, the term "sepsis" refers to a systemic inflammatory response syndrome to an infective process in which severe derangement of the immune system fails to prevent extensive ^λ spill over of inflammatory mediators from a local infection focus into the systemic circulation.

As used herein, the term "septic shock" refers to a consequence of sepsis in which the systemic inflammatory response leads to the failure of vital organ function (e.g., acute respiratory distress syndrome (ARDS)).

As used herein, the term "subject" broadly refers to any animal, including but not limited to, human and non-human animals (e.g., dogs, cats, cows, horses, sheep, poultry (e.g., chickens), fish, crustaceans, etc.). As used herein, the term "patient" typically refers to a human subject that is being treated for a disease or condition.

As used herein, the term "effective amount" refers to the amount of a composition sufficient to effect beneficial or desired results. An effective amount can be administered in one or more administrations, applications or dosages and is not intended to be limited to a particular formulation or administration route.

As used herein, the terms "administration" and "administering" refer to the act of giving a drug, prodrug, or other agent, or therapeutic treatment to a subject or in vivo, in vitro, or ex vivo cells, tissues, and organs. Exemplary routes of administration to the human body can be through space under the arachnoid membrane of the brain or spinal cord (intrathecal), the eyes (ophthalmic), mouth (oral), skin (topical or transdermal), nose (nasal), lungs (inhalant), oral mucosa (buccal), ear, rectal, vaginal, by injection (e.g., intravenously, subcutaneously, intratumorally, intraperitoneally, etc.) and the like.

As used herein, the terms "co-administration" and "co-administering" refer to the administration of at least two agent(s) (e.g., a pharmaceutical composition comprising beneficial bacteria, and/or additional therapeutics) or therapies to a subject. In some embodiments, the co-administration of two or more agents or therapies is concurrent. In other embodiments, a first agent/therapy is administered prior to a second agent/therapy. Those of skill in the art understand that the formulations and/or routes of administration of the various agents or therapies used may vary. The appropriate dosage for co-administration can be readily determined by one skilled in the art. In some embodiments, when agents or therapies are co-administered, the respective agents or therapies are administered at lower dosages than appropriate for their administration alone. Thus, co-administration is especially desirable in embodiments where the co-administration of the agents or therapies lowers the requisite dosage of a potentially harmful (e.g., toxic) agent(s), and/or when co-administration of two or more agents results in sensitization of a subject to beneficial effects of one of the agents via co-administration of the other agent.

As used herein, the term "treating," and linguistic variations thereof (e.g., "treat"), refers to the administration of a therapy or therapeutic to a subject suffering from a disease or condition with the goal or result of lessening the severity of the disease/condition, eliminating the disease/condition within the subject, and/or ameliorating symptoms of the

disease/condition within the subject.

As used herein, the term "preventing," and linguistic variations thereof (e.g., "prevent"), refers to the administration of a therapy or therapeutic to a subject, in a prophylactic manner, with the goal or result of (i) reducing the risk of the subject developing the disease or condition and/or (ii) reducing the severity of the disease or condition if the subject were to develop the disease or condition. The reduction in risk/severity may be total or partial, and in some embodiments may be measured by comparing the rate and/or severity of the disease/condition for a test population (e.g., subjects receiving the preventative treatment) relative to a control population.

As used herein, "risk of sepsis" refers to the likelihood that a subject will develop sepsis or related conditions (e.g., septic shock). A subject is at increased risk of sepsis when they have a bacterial, viral, fungal, or parasitic infection (e.g., at a wound, pneumonia, meningitis, etc.), and one or more ancillary risk factors, such as age (e.g., infants and elderly are at increased risk), impaired immune system, chronic or serious disease (e.g., diabetes, cancer), a metabolome or microbiome that promotes immune dysfunction, etc.

As used herein, the term "pharmaceutical composition" refers to the combination of an active agent with a carrier, inert or active, making the composition especially suitable for diagnostic or therapeutic use in vitro, in vivo or ex vivo.

The terms "pharmaceutically acceptable" or "pharmacologically acceptable," as used herein, refer to compositions that do not substantially produce adverse reactions, e.g., toxic, allergic, or immunological reactions, when administered to a subj ect.

As used herein, the term "pharmaceutically acceptable carrier" refers to any of the standard pharmaceutical carriers including, but not limited to, phosphate buffered saline solution, water, emulsions (e.g., such as an oil/water or water/oil emulsions), and various types of wetting agents, any and all solvents, dispersion media, coatings, sodium lauryl sulfate, isotonic and absorption delaying agents, disintigrants (e.g., potato starch or sodium starch glycolate), and the like. The compositions also can include stabilizers and

preservatives. For examples of carriers, stabilizers and adjuvants, see, e.g., Martin,

Remington's Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, Pa. (1975), incorporated herein by reference in its entirety.

As used herein, a "prebiotic" refers to an ingredient that allows specific changes, both in the composition and/or activity in the gastrointestinal microbiota that may (or may not) confer benefits upon the host. In some embodiments, a prebiotic is a comestible food or beverage or ingredient thereof. In some embodiments, a prebiotic is a selectively fermented ingredient. Prebiotics may include complex carbohydrates, amino acids, peptides, minerals, or other essential nutritional components for the survival of the bacterial composition.

Prebiotics include, but are not limited to, amino acids, biotin, fructooligosaccharide, galactooligosaccharides, hemicelluloses (e.g., arabinoxylan, xylan, xyloglucan, and glucomannan), inulin, chitin, lactulose, mannan oligosaccharides, oligofructose-enriched inulin, gums (e.g., guar gum, gum arabic and carregenaan), oligofructose, oligodextrose, tagatose, resistant maltodextrins (e.g., resistant starch), trans-galactooligosaccharide, pectins (e.g., xylogalactouronan, citrus pectin, apple pectin, and rhamnogalacturonan-I), dietary fibers (e.g., soy fiber, sugarbeet fiber, pea fiber, corn bran, and oat fiber) and

xylooligosaccharides.

As used herein, the term "small molecule" refers to molecules, whether naturally- occurring or artificially synthesized, that are non-polymeric (e.g., not proteins, polypeptides, nucleic acids, organic polymers, etc.) and not macromolecular (e.g., proteins, carbohydrates, lipids, nucleic acids). In some embodiments, small molecules are biologically active, in that they produce a local or systemic effect in animals, such as mammals, e.g., humans. In certain embodiments, small molecules are "gastrointestinally active" and induce specific changes in the composition and/or activity of gastrointestinal microbiota that may (or may not) confer benefits upon the host. Exemplary small molecules are antibiotics, signaling compounds, or others compounds that attenuates the virulence phenotype of pathogenic microbes.

As used herein, the term "antibody" refers to a whole antibody molecule or a fragment thereof (e.g., fragments such as Fab, scFv, Fab', Fc, Fv, diabodies, F(ab')2, etc.) unless specified otherwise. Embodiments referring to "an antibody" encompass multiple embodiments including "a whole antibody" and fragments of the antibody, which may alternatively be claimed or described using such language. The structure (e.g., light and heavy chains, constant and variable regions, complementarity determining regions (CDRs), etc.) and antigen/epitope specificity of antibodies and antibody fragments are well understood in the field.

As used herein, the term "microbe" refers to cellular prokaryotic and eukaryotic species from the domains Archaea, Bacteria, and Eukarya, the latter including yeast and filamentous fungi, protozoa, algae, or higher Protista, and encompasses both individual organisms and populations comprising any number of the organisms. The terms "microbial cells" and "microbes" are used interchangeably with the term "microorganism".

The term "prokaryotes" refers to cells that contain no nucleus or other cell organelles.

The prokaryotes are generally classified in one of two domains, the Bacteria and the Archaea. The definitive difference between organisms of the Archaea and Bacteria domains is based on fundamental differences in the nucleotide base sequence in the 16S ribosomal RNA. The terms "bacteria" and "bacterium" and "archaea" and "archaeon" refer to prokaryotic organisms of the domain Bacteria and Archaea in the three-domain system (see Woese CR, et al., Proc Natl Acad Sci U S A 1990, 87: 4576 - 79).

As used herein, the term "phylogenetic tree" refers to a graphical or schematic representation of the evolutionary relationships of one genetic sequence to another that is generated, for example, using a defined set of phylogenetic reconstruction algorithms (e.g. parsimony, maximum likelihood, or Bayesian). Nodes in the tree represent distinct ancestral sequences and the confidence of any node is provided, for example, by a bootstrap or Bayesian posterior probability, which measures branch uncertainty.

"rDNA", "rRNA", "16S-rDNA", "16S-rRNA", "16S", "16S sequencing", "16S- NGS", "18S", "18S-rRNA", "18S-rDNA", "18S sequencing", and "18S-NGS" refer to the nucleic acids that encode for the RNA subunits of the ribosome. rDNA refers to the gene that encodes the rRNA that comprises the RNA subunits. There are two RNA subunits in the ribosome termed the small subunit (SSU) and large subunit (LSU); the RNA genetic sequences (rRNA) of these subunits are related to the gene that encodes them (rDNA) by the genetic code. rDNA genes and their complementary RNA sequences are widely used for determination of the evolutionary relationships amount organisms as they are variable, yet sufficiently conserved to allow cross organism molecular comparisons. Typically 16S rDNA sequence (approximately 1542 nucleotides in length) of the 30S SSU is used for molecular- based taxonomic assignments of Prokaryotes and the 18S rDNA sequence (approximately 1869 nucleotides in length) of 40S SSU is used for Eukaryotes. 16S sequences are used for phylogenetic reconstruction as they are in general highly conserved, but contain specific hypervariable regions that harbor sufficient nucleotide diversity to differentiate genera and species of most bacteria. The "VI -V9 regions" of the 16S rRNA refers to the first through ninth hypervariable regions of the 16S rRNA gene that are used for genetic typing of bacterial samples. These regions in bacteria are defined by nucleotides 69-99, 137-242, 433- 497, 576-682, 822-879, 986-1043, 1117-1173, 1243-1294 and 1435-1465 respectively using numbering based on the E. coli system of nomenclature (Brosius et al, PNAS 75(10):4801- 4805 (1978); incorporated by reference in its entirety). In some embodiments, at least one of the VI, V2, V3, V4, V5, V6, V7, V8, and V9 regions are used to characterize an OTU. In one embodiment, the VI, V2, and V3 regions are used to characterize an OTU. In another embodiment, the V3, V4, and V5 regions are used to characterize an OTU. In another embodiment, the V4 region is used to characterize an OTU. A person of ordinary skill in the art can identify the specific hypervariable regions of a candidate 16S rRNA by comparing the candidate sequence in question to a reference sequence and identifying the hypervariable regions based on similarity to the reference hypervariable regions, or alternatively, one can employ Whole Genome Shotgun (WGS) sequence characterization of microbes or a microbial community.

As used herein, the term "operational taxonomic units" ("OTU") refers to a terminal leaf in a phylogenetic tree and is defined by a nucleic acid sequence, e.g., the entire genome, or a specific genetic sequence, and all sequences that share sequence identity to this nucleic acid sequence at the level of species. In some embodiments the specific genetic sequence may be the 16S sequence or a portion of the 16S sequence. In other embodiments, the entire genomes of two entities are sequenced and compared. In another embodiment, select regions such as multilocus sequence tags (MLST), specific genes, or sets of genes may be genetically compared. In some embodiments, OTUs that share≥97% average nucleotide identity across the entire 16S or some variable region of the 16S are considered the same OTU (see e.g. Claesson et al. 2010. Nucleic Acids Res 38: e200.; Konstantinidis et al. 2006. Philos Trans R Soc Lond B Biol Sci 361 : 1929-1940.; incorporated by reference in their entireties). In embodiments involving the complete genome, MLSTs, specific genes, or sets of genes OTUs that share≥95% average nucleotide identity are considered the same OTU (see e.g. Achtman M, and Wagner M. 2008. Nat. Rev. Microbiol. 6: 431-440.; Konstantinidis et al. 2006. Philos Trans R Soc LondB Biol Sci 361 : 1929-1940.; incorporated by reference in their entireties). OTUs are frequently defined by comparing sequences between organisms. Generally, sequences with less than 95% sequence identity are not considered to form part of the same OTU. OTUs may also be characterized by any combination of nucleotide markers or genes, in particular highly conserved genes (e.g., "house-keeping" genes), or a combination thereof. Such characterization employs, e.g., WGS data or a whole genome sequence.

The term "genus" is defined as a taxonomic group of related species according to the Taxonomic Outline of Bacteria and Archaea (Garrity et al. (2007) The Taxonomic Outline of Bacteria and Archaea. TOBA Release 7.7, March 2007. Michigan State University Board of Trustees).

The term "species" is defined as collection of closely related organisms with greater than 97% 16S ribosomal RNA sequence homology and greater than 70% genomic

hybridization and sufficiently different from all other organisms so as to be recognized as a distinct unit (e.g., an operational taxonomic unit).

The term "strain" as used herein in reference to a microorganism describes an isolate of a microorganism considered to be of the same species but with a unique genome and, if nucleotide changes are non-synonymous, a unique proteome differing from other strains of the same organism. Strains may differ in their non-chromosomal genetic complement.

Typically, strains are the result of isolation from a different host or at a different location and time, but multiple strains of the same organism may be isolated from the same host.

As used herein, the term "microbiota" refers to an assemblage of microorganisms localized to a distinct environment. Microbiota may include, for example, populations of various bacteria, eukaryotes (e.g., fungi), and/or archaea that inhabit a particular environment. For example, "gut microbiota," "vaginal microbiota," and "oral microbiota" refer to an assemblage of one or more species of microorganisms that are localized to, or found in, the gut, vagina, or mouth, respectively.

"Normal microbiota" refers to a population of microorganisms that localize in a particular environment in a normal, non-pathological state (e.g., a sample of gut microbiota from a subject without sepsis). A "normal microbiota" has normal membership and normal relative abundance.

"Abnormal microbiota" refers to a population of various microorganisms that localize in a particular environment in a subject suffering from or at risk of a pathological condition (e.g., a sample of gut microbiota from a subject with sepsis). Abnormal microbiota differs from normal microbiota in terms of identity (e.g., membership), absolute amount, or relative amount (e.g., relative abundance) of the various microbes.

As used herein, the term "commensal microbe" refers to a microorganism that is non- pathogenic to a host and is part of the normal microbiota of the host.

As used herein, the terms "microbial agent," "commensal microbial agent," and "probiotic" refer to compositions comprising a microbe or population of multiple different microbes for administration to a subject.

The term "biosynthetic pathway", also referred to as "metabolic pathway", refers to a set of anabolic or catabolic biochemical reactions for converting one chemical species into another. Gene products belong to the same "metabolic pathway" if they, in parallel or in series, act on the same substrate, produce the same product, or act on or produce a metabolic intermediate (e.g., a metabolite) between the same substrate and metabolite end product.

As used herein, the term "taxonomic unit" is a group of organisms that are considered similar enough to be treated as a separate unit. A taxonomic unit may comprise, e.g., a class, family, genus, species, or population within a species (e.g., strain), but is not limited as such.

As used herein, a "colony -forming unit" ("CFU") is used as a measure of viable microorganisms in a sample. A CFU is an individual viable cell capable of forming on a solid medium a visible colony whose individual cells are derived by cell division from one parental cell.

As used herein, the term "relative abundance" relates to the abundance of

microorganisms of a particular taxonomic unit or OTU in a test biological sample compared to the abundance of microorganisms of the corresponding taxonomic unit or OTU in one or more non-diseased control samples. The "relative abundance" may be reflected in e.g., the number of isolated species corresponding to a taxonomic unit or OTU or the degree to which a biomarker specific for the taxonomic unit or OTU is present or expressed in a given sample. The relative abundance of a particular taxonomic unit or OTU in a sample can be determined using culture-based methods or non-culture-based methods well known in the art. Non- culture based methods include sequence analysis of amplified polynucleotides specific for a taxonomic unit or OTU or a comparison of proteomics-based profiles in a sample reflecting the number and degree of polypeptide-based, lipid-based, polysaccharide-based or carbohydrate-based biomarkers characteristic of one or more taxonomic units or OTUs present in the samples. Relative abundance or abundance of a taxon or OTU can be calculated with reference to all taxa/OTUs detected, or with reference to some set of invariant taxa/OTUs.

Methods for profiling the relative abundances of microbial taxa in biological samples, including biological samples of gut microbiota, are well known in the art. Suitable methods may be sequencing-based or array-based. For example, the microbial component of a gut microbiota sample is characterized by sequencing a nucleic acid suitable for taxonomic classification and assigning the sequencing reads to operational taxonomic units (OTUs) with a defined (e.g., > 97%) nucleotide sequence identity to a database of annotated and representative sequences. An example of such a database is Greengenes version of May 2013; however any suitable database may be used. After OTUs are defined, a representative sequence from each OTU can be selected and compared to a reference set. If a match is identified in the reference set, that OTU can be given an identity. Relative abundance of a bacterial taxon may be defined by the number of sequencing reads that can be unambiguously assigned to each taxon after adjusting for genome uniqueness. Other methods of profiling the relative abundances of microbial taxa in biological samples are known within the field and within the scope herein.

In some embodiments, a suitable nucleic acid for taxonomic classification is universally distributed among the gut microbial population being queried allowing for the analysis of phylogenetic relationships among distant taxa, and has both a conserved region and at least one region subject to variation. The presence of at least one variable region allows sufficient diversification to provide a tool for classification, while the presence of conserved regions enables the design of suitable primers for amplification (if needed) and/or probes for hybridization for various taxa at different taxonomic levels ranging from individual strains to whole phyla. While any suitable nucleic acid known in the art may be used, one skilled in the art will appreciate that selection of a nucleic acid or region of a nucleic acid to amplify may differ by environment. In some embodiments, a nucleic acid queried is a small subunit ribosomal RNA gene. For bacterial and archaeal populations, at least the VI, V2, V3, V4, V5, V6, V7, V8, and/or V9 regions of the 16S rRNA gene are suitable, though other suitable regions are known in the art. Guidance for selecting a suitable 16S rRNA region to amplify can be found throughout the art, including Guo et al. PLOS One 8(10) e76185, 2013; Soergel DAW et al. ISME Journal 6: 1440, 2012; and Hamady M et al. Genome Res. 19: 1 141 , 2009, each hereby incorporated by reference in its entirety.

DETAILED DESCRIPTION

Provided herein are compositions (e.g., probiotic, therapeutics, pharmaceutical, etc.) comprising one or more strains of bacteria (e.g., selected from the strains listed in Tables 1 -6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, and/or Bacteroidaceae. In particular, provided herein are compositions comprising bacteria from one or more of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus and methods of use thereof for inducing immune system maintenance and/or rescuing humans and animals from infections, including sepsis.

Experiments conducted during development of embodiments herein have

demonstrated that restoration of the gut microbiota, for example, with fecal microbiota transplant (FMT) protects against advanced-stage polymicrobial sepsis. For example, using a murine model of lethal sepsis that involves the introduction of a four-member pathogen community (PC) into the cecum of surgically stressed mice, septic mice were treated with FMT consisting of feces from healthy mice, which led to a drastic increase in survival compared to mice that were treated with heat-inactivated FMT. The protective effect of FMT was associated with re-establishment of gut microbiota diversity and reinstatement of immune homeostasis. Reiterative studies were conducted during development of

embodiments herein in which the PC was administered systemically through injection into the peritoneum (IP) and FMT was administered to the gut with similar results on mortality. Comparative transcriptomics indicates that in both gut-derived sepsis and systemic (IP) sepsis, the FMT rescue effect involved maintenance of innate immune signaling pathways. The bacterial families S24-7 (Prevotella-like), Rikenellaceae, Prevotellaceae,

Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and

Bacteroidaceae are significantly associated with immune system activation and stable immune-inflammation response (e.g., to sepsis). The experiments conducted during development of embodiments herein indicate that a combination of bacterial strains (e.g., 1 strain, 2 strains, 3 strains, 4 strains, 5 strains, 6 strains, 7 strains, 8 strains, 9 strains, 10 strains, 15 strains, 20 strains, 30 strains, 40 strains, 50 strains, 75 strains, 100 strains, 200 strains, 500 strains, 1000 strains, or more or ranges therebetween (e.g., 5-1 1 strains, 7-9 strains, etc.)) belonging to the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus are efficacious in inducing immune system maintenance and preventing and/or rescuing animals from serious infections, including sepsis. In some embodiments, the combination of bacterial strains comprises, or consists of, bacterial strains selected from Table 1, Table 2, Table 3, Table 4, Table 5, and/or Table 6.

Experiments conducted during development of embodiments herein indicated that certain strains and species of bacteria, for example from the families Prevotellaceae,

Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae; the genera Prevotella, Paraprevotella, Alistipes, Bacteroides,

Lactobacillus, Parabacteroides, and/or Ruminococcus; and/or the strains listed in Tables 1-6 are important for enabling stabilization of immune response, particularly to serious infections including sepsis. In some embodiments, organisms belonging to the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus (e.g., bacteria of the strains listed in Tables 1-6) modulate immune stabilization and/or response to infections, including sepsis.

Accordingly, in some embodiments, the present technology provides compositions and kits, e.g., for administration to a subject. In some embodiments, compositions comprise one or more bacterial species or strains (e.g., selected from the strains listed in Tables 1-6) of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae,

Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus. Embodiments are not limited to a particular one or more bacterial species; although compositions and kits comprising one or more bacterial strains (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, or more or ranges therebetween) of the strains listed in Tables 1-6 are within the scope herein.

In some embodiments, compositions/kits comprise a single species of bacteria. In other embodiments, the compositions/kits comprise two or more species of bacteria, e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, 100, 500, 1000 or more, or ranges therebetween species of bacteria. In one embodiment, compositions/kits comprise no more than 20 species of bacteria, e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 species of bacteria. In some embodiments, compositions/kits comprise bacteria of one or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, or more or ranges therebetween) strains listed in Table 1, Table 2, Table 3, Table 4,Table 5 and/or Table 6. In some embodiments, methods of administering such compositions/kits are provided.

In some embodiments, compositions/kits comprise a single OTU. In some embodiments, compositions/kits comprise two or more OTUs (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, 100, 500, 1000 or more, or ranges therebetween). In some embodiments, each OTU independently characterized by, for example, at least 95%, 96%, 97%, 98%, 99% 100% sequence identity a reference sequence (e.g., a segment of 16S RNA) for the OTU (e.g., species, genus, etc.).

In some embodiments, compositions comprise one or more bacteria species/strains from the family Prevotellaceae (e.g., bacteria of the species or strains listed in Table 1). In some embodiments, the Prevotellaceae bacteria are of one or more genera selected from Prevotella, Alloprevotella, Hallella and/or Paraprevotella. In some embodiments, compositions comprise one or more Prevotellaceae bacteria from the species and strains listed in Table 1.

In some embodiments, compositions comprise one or more bacteria species/strains from the genus Prevotella. In some embodiments, the Prevotella bacteria are of one or more species selected from Prevotella albensis, Prevotella amnii, Prevotella bergensis, Prevotella bivia, Prevotella brevis, Prevotella bryantii, Prevotella buccae, Prevotella buccalis, Prevotella copri, Prevotella dentalis, Prevotella denticola, Prevotella disiens, Prevotella histicola,

Prevotella intermedia, Prevotella maculosa, Prevotella marshii, Prevotella melaninogenica, Prevotella micans, Prevotella multiformis, Prevotella nigrescens, Prevotella oralis, Prevotella oris, Prevotella oulorum, Prevotella pallens, Prevotella salivae, Prevotella stercorea, Prevotella tannerae, Prevotella timonensis, Prevotella veroralis.

In some embodiments, compositions comprise one or more bacteria species/strains from the genus Paraprevotella. In some embodiments, the Paraprevotella bacteria are of one or more species selected from Paraprevotella xylaniphila and Paraprevotella clara.

In some embodiments, compositions comprise one or more bacteria species/strains from the family Porphyromonadaceae. In some embodiments, the Porphyromonadaceae bacteria are of one or more genera selected from Porphyromonas, Dysgonomonas, and/or Macellibacteroides.

In some embodiments, compositions comprise one or more bacteria species/strains from the family Lactobacillaceae (e.g., bacteria of the species or strains listed in Table 3). In some embodiments, the Lactobacillaceae bacteria are of one or more genera selected from Lactobacillus, Paralactobacillus, Pediococcus, and/or Sharpea. In some embodiments, compositions comprise one or more Lactobacillaceae bacteria from the species and strains listed in Table 3.

In some embodiments, compositions comprise one or more bacteria species/strains from the genus Lactobacillus. In some embodiments, the Lactobacillus bacteria are of one or more species selected from L. acetotolerans, L. acidifarinae, L. acidipiscis, L. acidophilus, L. agilis, L. algidus, L. alimentarius, L. amylolyticus, L. amylophilus, L. amylotrophicus, L. amylovorus, L. animalis, L. antri, L. apodemi, L. aviaries, L. bifermentans, L. brevis, L.

buchneri, L. camelliae, L. casei, L. catenaformis, L. ceti, L. coleohominis, L. collinoides, L. composti, L. concavus, L. coryniformis, L. crispatus, L. crustorum, L. curvatus, L.

delbrueckii subsp. Bulgaricus, L. delbrueckii subsp. Delbrueckii, L. delbrueckii subsp. Lactis,

L. dextrinicus, L. diolivorans, L. equi, L. equigenerosi, L. farraginis, L. farciminis, L.

fermentum, L. fomicalis, L. fructivorans, L. frumenti, L. fuchuensis, L. gallinarum, L. gasseri,

L. gastricus, L. ghanensis, L. graminis, L. hammesii, L. hamster, L. harbinensis, L.

hayakitensis, L. helveticus, L. hilgardii, L. homohiochii, L. iners, L. ingluviei, L. intestinalis,

L. jensenii, L. johnsonii, L. kalixensis, L. kefiranofaciens, L. kefiri, L. kimchi, L. kitasatonis,

L. kunkeei, L. leichmannii, L. lindneri, L. malefermentans, L. mali, L. manihotivorans, L. mindensis, L. mucosae, L. murinus, L. nagelii, L. namurensis, L. nantensis, L.

oligofermentans, L. oris, L. panis, L. pantheris, L. parabrevis, L. parabuchneri, L. paracasei, L. paracollinoides, L. arafarraginis, L. parakefiri, L. paralimentarius, L. paraplantarum, L. pentosus, L. perolens, L. plantarum, L. pontis, L. protectus, L. psittaci, L. rennini, L. reuteri,

L. rhamnosus, L. rimae, L. rogosae, L. rossiae, L. ruminis, L. saerimneri, L. sakei, L.

salivarius, L. sanfranciscensis, L. satsumensis, L. secaliphilus, L. sharpeae, L. siliginis, L. spicheri, L. suebicus, L. thailandensis, L. ultunensis, L. vaccinostercus, L. vaginalis, L.

versmoldensis, L. vini, L. vitulinus, L. zeae, and/or L. zymae.

In some embodiments, compositions comprise one or more bacteria species/strains from the family Bacteroidaceae (e.g., bacteria of the species or strains listed in Table 2). In some embodiments, the Bacteroidaceae bacteria are of one or more genera selected from

Bacteroides, Acetofilamentum, Acetomicrobium, Acetothermus, and/or Anaerorhabdus. In some embodiments, compositions comprise one or more Bacteroidaceae bacteria from the species and strains listed in Table 2.

In some embodiments, compositions comprise one or more bacteria species/strains from the genus Bacteroides. In some embodiments, the Bacteroides bacteria are of one or more species selected from B. acidifaciens, B. gracilis, B. fragilis, B. oris, B. ovatus, B. putredinis, B. pyogenes, B. stercoris, B. suis, B. tectus, B. thetaiotaomicron, and.or B.

vulgatus.

In some embodiments, compositions comprise one or more bacteria species/strains from the genus Parabacteroides. In some embodiments, the Parabacteroides bacteria are of one or more species selected from Parabacteroides distasonis, Parabacteroides goldsteinii, and/or Parabacteroides merdae.

In some embodiments, compositions comprise one or more bacteria species/strains from the family Ruminococcaceae (e.g., bacteria of the species or strains listed in Table 4). In some embodiments, the Ruminococcaceae bacteria are of the genus Ruminococcus. In some embodiments, compositions comprise one or more Ruminococcaceae bacteria from the species and strains listed in Table 4.

In some embodiments, compositions comprise one or more bacteria species/strains from the family Rikenellaceae. In some embodiments, the Rikenellaceae bacteria are of one or more genera selected from Rikenella, Alistipes, and/or Anaerocella. In some embodiments, compositions comprise one or more Rikenellaceae bacteria from the species and strains listed in Table 5.

In some embodiments, compositions comprise one or more bacteria species/strains from the family Lachnospiraceae (e.g., bacteria of the species or strains listed in Table 6). In some embodiments, the Lachnospiraceae bacteria are of one or more genera selected from Abyssivirga, Acetatifactor, Acetitomaculum, Agathobacter, Anaerostipes, Butyrivibrio, Catonella, Cellulosilyticum, Coprococcus, Dorea, Hespellia Johnsonella,

Lachnoanaerobaculum, Lachnobacterium, Lachnospira, Marvinbryantia, Mobilitalea, Moryella, Oribacterium, Parasporobacterium, Pseudobutyrivibrio, Robinsoniella, Roseburia, Shuttleworthia, Sporobacterium, Stomatobaculum, and/or Syntrophococcus. In some embodiments, compositions comprise one or more Lachnospiraceae bacteria from the species and strains listed in Table 6.

In some embodiments, compositions comprise one or more bacteria species/strains from the genus Ruminococcus. In some embodiments, the Ruminococcus bacteria are of one or more species selected from Ruminococcus albus, Ruminococcus bromii, Ruminococcus callidus, Ruminococcus flavefaciens, Ruminococcus gauvreauii (reclassified in the

Lachnospiraceae family), Ruminococcus gnavus (reclassified in the Lachnospiraceae family), Ruminococcus lactaris (reclassified in the Lachnospiraceae family), Ruminococcus obeum (reclassified in the Lachnospiraceae family), and/or Ruminococcus torques (reclassified in the Lachnospiraceae family). In some embodiments, compositions comprise one or more Ruminococcus bacteria from the species and strains listed in Table 4.

In some embodiments, compositions comprise one or more additional components (e.g., including but not limited to, one or more additional additive(s) selected from the group consisting of an energy substrate, a source of nitrogen, phosphorus or iron [Fe(II) and/or Fe(III)], a mineral, a vitamin, or combinations thereof).

In some embodiments, in addition to bacteria selected from the families

Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus, a composition or kit comprises one or more beneficial and/or commensal bacteria selected from other genera and/or taxa.

In some embodiments, bacteria are vegetative cells, freeze-dried cells, where possible spores, etc. Freeze-dried bacteria can be stored for several years with maintained viability. In certain applications, freeze-dried bacteria are sensitive to humidity. One way of protecting the bacterial cells is to store them in oil. The freeze dried bacterial cells can be mixed directly with a suitable oil, or alternately the bacterial cell solution can be mixed with an oil and freeze dried together, leaving the bacterial cells completely immersed in oil. Suitable oils may be edible oils such as olive oil, rapeseed oil which is prepared conventionally or cold- pressed, sunflower oil, soy oil, maize oil, cotton-seed oil, peanut oil, sesame oil, cereal germ oil such as wheat germ oil, grape kernel oil, palm oil and palm kernel oil, linseed oil. The viability of freeze-dried bacteria in oil is maintained for at least nine months. Optionally live cells can be added to one of the above oils and stored.

In some embodiments, compositions are added to nutraceuticals, food products, or foods. In some embodiments, to give the composition or nutraceutical a pleasant taste, flavoring substances such as for example mints, fruit juices, licorice, Stevia rebaudiana, steviosides or other calorie free sweeteners, rebaudioside A, essential oils like eucalyptus oil, or menthol can optionally be included in compositions of embodiments of the present invention.

In some composition embodiments, compositions are formulated in pharmaceutical compositions. The bacteria of embodiments herein may be administered alone or in combination with pharmaceutically acceptable carriers or diluents, and such administration may be carried out in single or multiple doses as described herein. Compositions may, for example, be in the form of tablets, resolvable tablets, capsules, bolus, drench, pills sachets, vials, hard or soft capsules, aqueous or oily suspensions, aqueous or oily solutions, emulsions, powders, granules, syrups, elixirs, lozenges, reconstitutable powders, liquid preparations, creams, troches, hard candies, sprays, chewing-gums, creams, salves, jellies, gels, pastes, toothpastes, rinses, dental floss and tooth-picks, liquid aerosols, dry powder formulations, HFA aerosols or organic or inorganic acid addition salts.

The pharmaceutical compositions of embodiments of the invention may be in a form suitable for, e.g., rectal, oral, topical, buccal administration. Depending upon the disorder and patient to be treated and the route of administration, the compositions may be administered at varying doses.

In some embodiments, one or more bacteria strains or species from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus (e.g., alone or with other active components) are formulated in pharmaceutical compositions for rectal administration. Such formulations include enemas, rectal gels, rectal foams, rectal aerosols, suppositories, jelly suppositories, or retention enemas, containing conventional suppository bases such as cocoa butter or other glycerides, as well as synthetic polymers such as polyvinylpyrrolidone, PEG, Pi-PEG, and the like. In suppository forms of the compositions, a low-melting wax such as, but not limited to, a mixture of fatty acid glycerides, optionally in combination with cocoa butter is first melted.

In some embodiments, one or more bacteria strains or species (e.g., the bacterial strains and/or species listed in Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides,

Lactobacillus, Parabacteroides, and/or Ruminococcus (e.g., alone or with other active components) are formulated in pharmaceutical compositions for oral administration. Oral dosage forms include push fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. In specific embodiments, push fit capsules contain the active ingredients in admixture with one or more filler. Fillers include, by way of example only, lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In other embodiments, soft capsules, contain one or more active compound that is dissolved or suspended in a suitable liquid. Suitable liquids include, by way of example only, one or more fatty oil, liquid paraffin, or liquid polyethylene glycol. In addition, stabilizers are optionally added.

In some embodiments, the bacterial formulation comprises at least 1 ^χΐθ⁴ CFU (e.g., lxlO⁴ CFU, 2xl0⁴ CFU, 5xl0⁴ CFU, lxlO⁵ CFU, 2xl0⁵ CFU, 5xl0⁵ CFU, lxlO⁶ CFU, 2xl0⁶ CFU, 5xl0⁶ CFU, lxlO⁷ CFU, 2xl0⁷ CFU, 5xl0⁷ CFU, lxlO⁸ CFU, 2xl0⁸ CFU, 5xl0⁸ CFU, lxlO⁹ CFU, 2xl0⁹ CFU, 5xl0⁹ CFU, lxlO¹⁰ CFU, 2xl0¹⁰ CFU, 5xl0¹⁰ CFU, lxlO¹¹ CFU, 2xlOⁿ CFU, 5xlOⁿ CFU, lxlO¹² CFU, 2xl0¹² CFU, 5xl0¹² CFU, or more or ranges there between) bacteria (e.g., species or strains selected from Tables 1-6) from the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae,

Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus (e.g., from a single classification (e.g., strain, species, genus, etc.), alone or with other active components. In some embodiments, the bacterial formulation is administered to the subject in two or more doses (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more, or ranges there between). In some embodiments, the administration of doses are separated by at least 1 day (e.g., 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 2 weeks, 3 weeks, 4 weeks, or ranges there between).

For oral or buccal administration, bacteria of embodiments of the present invention may be combined with various excipients. Solid pharmaceutical preparations for oral administration often include binding agents (for example syrups, acacia, gelatin, tragacanth, polyvinylpyrrolidone, sodium lauryl sulphate, pregelatinized maize starch, hydroxypropyl methylcellulose, starches, modified starches, gum acacia, gum tragacanth, guar gum, pectin, wax binders, microcrystalline cellulose, methylcellulose, carboxymethylcellulose, hydroxypropyl methylcellulose, hydroxy ethyl cellulose, hydroxypropyl cellulose,

copolyvidone and sodium alginate), disintegrants (such as starch and preferably com, potato or tapioca starch, alginic acid and certain complex silicates, polyvinylpyrrolidone, gelatin, acacia, sodium starch gly collate, microcrystalline cellulose, crosscarmellose sodium, crospovidone, hydroxypropyl methylcellulose and hydroxypropyl cellulose), lubricating agents (such as magnesium stearate, sodium lauryl sulfate, talc, silica polyethylene glycol waxes, stearic acid, palmitic acid, calcium stearate, carnuba wax, hydrogenated vegetable oils, mineral oils, polyethylene glycols and sodium stearyl fumarate) and fillers (including high molecular weight polyethylene glycols, lactose, calcium phosphate, glycine magnesium stearate, starch, rice flour, chalk, gelatin, microcrystalline cellulose, calcium sulphate, and lactitol). Such preparations may also include preservative agents and anti-oxidants. Liquid compositions for oral administration may be in the form of, for example, emulsions, syrups, or elixirs, or may be presented as a dry product for reconstitution with water or other suitable vehicle before use. Such liquid compositions may contain

conventional additives such as suspending agents (e.g. syrup, methyl cellulose, hydrogenated edible fats, gelatin, hydroxyalkylcelluloses, carboxymethylcellulose, aluminium stearate gel, hydrogenated edible fats) emulsifying agents (e.g. lecithin, sorbitan monooleate, or acacia), aqueous or non-aqueous vehicles (including edible oils, e.g. almond oil, fractionated coconut oil) oily esters (for example esters of glycerine, propylene glycol, polyethylene glycol or ethyl alcohol), glycerine, water or normal saline; preservatives (e.g. methyl or propyl p- hydroxy benzoate or sorbic acid) and conventional flavoring, preservative, sweetening or coloring agents. Diluents such as water, ethanol, propylene glycol, glycerin and combinations thereof may also be included.

Other suitable fillers, binders, disintegrants, lubricants and additional excipients are well known to a person skilled in the art.

In some embodiments, microbes are spray-dried. In other embodiments, microbes are suspended in an oil phase and are encased by at least one protective layer, which is water- soluble (water-soluble derivatives of cellulose or starch, gums or pectins; See e.g., EP 0 180 743, herein incorporated by reference in its entirety).

In some embodiments, the compositions, kits, and methods herein are utilized in addition to other treatments and/or prophylactics for sepsis and/or infection. In some embodiments, pharmaceutical compositions, and/or kits comprise antibiotics to inhibit the growth of microorganisms. For example, the antibiotic may inhibit cell wall synthesis, protein synthesis, nucleic acid synthesis, or alter cell membrane function. Classes of antibiotics that find use in conjunction with the other embodiments herein include, but are not limited to, macrolides (i.e., erythromycin), penicillins (i.e., nafcillin), cephalosporins (i.e., cefazolin), carbepenems (i.e., imipenem, aztreonam), other beta-lactam antibiotics, beta- lactam inhibitors (i.e., sulbactam), oxalines (i.e. linezolid), aminoglycosides (i.e., gentamicin), chloramphenicol, sulfonamides (i.e., sulfamethoxazole), glycopeptides (i.e., vancomycin), quinolones (i.e., ciprofloxacin), tetracyclines (i.e., minocycline), fusidic acid, trimethoprim, metronidazole, clindamycin, mupirocin, rifamycins (i.e., rifampin), streptogramins (i.e., quinupristin and dalfopristin) lipoprotein (i.e., daptomycin), polyenes (i.e., amphotericin B), azoles (i.e., fluconazole), and echinocandins (i.e., caspofungin acetate). Examples of specific antibiotics that can be used include, but are not limited to, erythromycin, nafcillin, cefazolin, imipenem, aztreonam, gentamicin, sulfamethoxazole, vancomycin, ciprofloxacin, trimethoprim, rifampin, metronidazole, clindamycin, teicoplanin, mupirocin, azithromycin, clarithromycin, ofloxacin, lomefloxacin, norfloxacin, nalidixic acid, sparfloxacin, pefloxacin, amifloxacin, gatifloxacin, moxifloxacin, gemifloxacin, enoxacin, fleroxacin, minocycline, linezolid, temafloxacin, tosufloxacin, clinafloxacin, sulbactam, clavulanic acid, amphotericin B, fluconazole, itraconazole, ketoconazole, and nystatin.

In some embodiments, pharmaceutical compositions, and/or kits comprise small molecules to attenuate the virulence response of microorganisms. For example, the small molecule may inhibit the expression of collagenases and proteolytic activities related to virulence, such as activation of intestinal tissue matrix metalloprotease-9 (MMP9). Such small molecules that attenuate the virulence response include, but are not limited to phosphorylated PEG (e.g. Pi-PEG15-20).

In some embodiments, anti-viral agents are used in combination the compositions herein to treat and/or prevent a sepsis or underlying conditions. Such anti-viral agents include, but are not limited to protease inhibitors (e.g., saquinavir, ritonavir, amprenavir), reverse transcriptase inhibitors (e.g., azidothymidine (AZT), lamioridine (3TC), dideoxyinosine

(ddl)), dideoxycytidine (ddC), zidovudine), nucleoside analogs (e.g., acyclovir, penciclovir).

Other anti-sepsis agents include, but are not limited to Drotrecogin alfa (activated). Anti-inflammatory agents, such as non-steroidal anti -inflammatory agents (e.g., naproxen, ibuprofen, celeoxib) and steroidal anti-inflammatory agents (e.g., glucocorticoids) also find use in some embodiments herein.

In some embodiments, the present technology provides kits, pharmaceutical compositions, or other delivery systems for use in treatment or prevention of sepsis in a subject. The kit may include any and all components necessary, useful or sufficient for research or therapeutic uses including, but not limited to, one or more microbes (e.g., bacteria selected from the species and/or strains listed in Tables 1 -6) from the families

Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and Ruminococcus, and other components, such as pharmaceutical carriers, and additional components useful, necessary or sufficient for use in treatment or prevention of sepsis. In some embodiments, the kits provide a sub-set of the required components, wherein it is expected that the user will supply the remaining components. In some embodiments, the kits comprise two or more separate containers wherein each container houses a subset of the components to be delivered. Optionally, compositions and kits comprise other active components in order to achieve desired therapeutic effects.

In some embodiments, compositions and kits provided herein (e.g., comprising Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae and/or Bacteroidaceae, Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus bacteria) are administered once to a subject in need thereof (e.g., a subject suffering from or at risk of sepsis).

In some embodiments, compositions comprise prebiotic compounds such as carbohydrate compounds selected from the group consisting of inulin, fructooligosaccharide (FOS), short-chain fructooligosaccharide (short chain FOS), galacto-oligosaccharide (GOS), xylooligosaccharide (XOS), glangliosides, partially hydrolysed guar gum (PHGG) acacia gum, soybean-gum, apple extract, lactowolfberry, wolfberry extracts or mixture thereof.

Other carbohydrates may be present such as a second carbohydrate acting in synergy with the first carbohydrate and that is selected from the group consisting of xylooligosaccharide (XOS), gum, acacia gum, starch, partially hydrolysed guar gum or mixture thereof. The carbohydrate or carbohydrates may be present at about 1 g to 20g or 1 % to 80% or 20% to 60% in the daily doses of the composition. Alternatively, the carbohydrates are present at 10% to 80% of the dry composition.

The daily doses of carbohydrates, and all other compounds administered with the probiotics comply with published safety guidelines and regulatory requirements. This is particularly important with respect to the administration to newborn babies.

In some embodiments, a nutritional composition preferably comprises a source of protein. Dietary protein is preferred as a source of protein. The dietary protein may be any suitable dietary protein, for example animal proteins, vegetable proteins (such as soy proteins, wheat proteins, rice proteins or pea proteins), a mixture of free amino acids, or a combination thereof. The composition may also comprise a source of carbohydrates, nitrogen, phosphorus, essential co-factor, minerals, and/or a source of fat.

In some embodiments, compositions are administered on an ongoing, recurrent, or repeat basis (e.g., multiple times a day, once a day, once every 2, 3, 4, 5, or 6 days, once a week, etc.) for a period of time (e.g., multiple days, months, or weeks). Suitable dosages and dosing schedules are determined by one of skill in the art using suitable methods. In some embodiments, compositions are administered prior to an event (e.g., surgery) in which the subject may be at greater risk of infection or sepsis. In some embodiments, the combination of Prevotellaceae, Rikenellaceae,

Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus bacteria strains (e.g., selected from the species and/or strains listed in Tables 1-6) is selected to mimic the healthy microbiota of a non-septic subject. In some

embodiments, the combination Prevotellaceae, Rikenellaceae, Porphyromonadaceae,

Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella,

Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus bacteria strains (e.g., selected from the species and/or strains listed in Tables 1-6) is selected to generate a healthy microbiota of a non-septic subject. In some embodiments, the combination of Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus bacteria strains is selected to generate the healthy metabolite makeup of non-septic subject.

In some embodiments, methods are provided herein for (1) preventing development of infections that can lead to sepsis in a subject, (2) preventing the development of sepsis from an infection or potential infection (e.g., should a subject become infected) and/or (3) treating a subj ect suffering from infections that can lead to sepsis. In some embodiments, a subject suffers from, is at risk of suffering, or is at increased risk of suffering from sepsis. In some embodiments, the sepsis or risk of sepsis is caused by a bacterial, viral, or fungal infection, such as pneumonia, an abdominal infection, a kidney infection, or a bloodstream infection (e.g., bacteremia). In some embodiments, the subj ect exhibits one or more of: abnormal body temperature (e.g., above 101 °F (38.3 °C) or below 96.8 °F (36 °C) for a human subject), increased heart rate (e.g., above 90 beats per minute for a human subject), and/or increased respiratory rate (e.g., higher than 20 breaths a minute for a human subject). In some embodiments, the subject suffers from or is at risk of severe sepsis, for example, as indicated by one or more of significantly decreased urine output, abrupt change in mental status, decrease in platelet count, difficulty breathing, abnormal heart pumping function, and/or abdominal pain. In some embodiments, the subject suffers from or is at risk of septic shock, as evidenced by one or more of the above symptoms in addition to extremely low blood pressure that doesn't adequately respond to simple fluid replacement. In some embodiments, a subject is treated by the methods herein to prevent development of one or more of the aforementioned symptoms or conditions (e.g., sepsis, severe sepsis, septic shock, etc.). In some embodiments, a subject is treated by the methods herein to alleviate one or more of the aforementioned symptoms and/or treat one for the aforementioned conditions (e.g., sepsis, severe sepsis, septic shock, etc.). In some embodiments, a composition comprising bacteria (e.g., selected from the species and/or strains listed in Tables 1-6) of the families

Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae, and/or the genera Prevotella, Paraprevotella,

Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus is administered to a subject or patient in a pharmaceutically effective amount.

The dosage amount and frequency are selected to create an effective level of the bacteria described herein without substantially harmful effects. When administered (e.g., orally, rectally, etc.), the dosage will generally comprise at least 1 ^χΐθ⁴ CFU per dose or per day (e.g., lxlO⁴ CFU, 2xl0⁴ CFU, 5xl0⁴ CFU, lxlO⁵ CFU, 2xl0⁵ CFU, 5xl0⁵ CFU, lxlO⁶ CFU, 2xl0⁶ CFU, 5xl0⁶ CFU, lxlO⁷ CFU, 2xl0⁷ CFU, 5xl0⁷ CFU, lxlO⁸ CFU, 2xl0⁸ CFU, 5xl0⁸ CFU, lxlO⁹ CFU, 2xl0⁹ CFU, 5xl0⁹ CFU, lxlO¹⁰ CFU, 2xl0¹⁰ CFU, 5xl0¹⁰ CFU, lxlO¹¹ CFU, 2xlOⁿ CFU, 5xl0ⁿ CFU, lxlO¹² CFU, 2xl0¹² CFU, 5xl0¹² CFU per dose or per day, or more per dose or per day, including ranges there between) of Prevotellaceae,

Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus,

Parabacteroides, and/or Ruminococcus bacteria (e.g., species or strains of bacteria selected from those listed in Tables 1-6).

Methods of administering a composition described herein include, without limitation, administration in oral, intranasal, topical, sublingual, rectal, and vaginal forms.

In some embodiments, a single dose of a composition comprising Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus,

Parabacteroides, and/or Ruminococcus bacteria (e.g., species or strains of bacteria selected from those listed in Tables 1-6) is administered to a subject. In other embodiments, multiple doses are administered over two or more time points, separated by hours, days, weeks, etc. In some embodiments, a composition comprising Prevotellaceae, Rikenellaceae,

Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or

Ruminococcus bacteria (e.g., an effective level of bacteria) is administered over a long period of time (e.g., chronically), for example, for a period of months or years (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or more months or years; for the subject's lifetime). In such embodiments, a composition comprising Prevotellaceae, Rikenellaceae, Po hyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella,

Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus bacteria (e.g., species or strains of bacteria selected from those listed in Tables 1-6) (e.g., an effective level) may be taken on a regular scheduled basis (e.g., daily, weekly, etc.) for the duration of the extended period.

In some embodiments, subjects are tested to determine their microbiota (e.g., to if the microbiota places the subject at risk of sepsis). Exemplary diagnostic methods are described herein. In some embodiments, intact bacteria are detected (e.g., by detecting surface polypeptides or markers). In other embodiments, bacteria are lysed and nucleic acids or proteins (e.g., corresponding to genes specific to the species of bacteria) are detected. In some embodiments, bacteria are identified using detection reagents (e.g., a probe, a microarray, e.g., an amplification primer) that specifically interact with a nucleic acid that identifies a particular species of bacteria (e.g., beneficial bacteria, pathogenic bacteria, commensal bacteria, non-commensal bacteria; bacteria of the genera Prevotella,

Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides and/or Ruminococcus, etc.).

Some embodiments comprise use of nucleic acid sequencing to detect, quantify, and/or identify gut microbiota. The term "sequencing," as used herein, refers to a method by which the identity of at least 10 consecutive nucleotides (e.g., the identity of at least 20, at least 50, at least 100, or at least 200 or more consecutive nucleotides) of a polynucleotide are obtained. The term "next-generation sequencing" refers to the so-called parallelized sequencing-by-synthesis or sequencing-by-ligation platforms currently employed by Illumina, Life Technologies, and Roche, etc. Next-generation sequencing methods may also include nanopore sequencing methods or electronic-detection based methods such as Ion Torrent technology commercialized by Life Technologies.

Some embodiments of the technology comprise acquiring a gut microbiota sample from a subject. As used herein, "gut microbiota sample" refers to a biological sample comprising a plurality of heterogeneous nucleic acids produced by a subject's gut microbiota. Fecal samples are commonly used in the art to sample gut microbiota, but samples can also be taken from the small intestine (e.g., duodenum, jeunum, ileum) or the large intestine (e.g., cecum, colon, rectum, anal canal). Methods for obtaining a fecal sample from a subject are known in the art and include, but are not limited to, rectal swab and stool collection. Suitable fecal samples may be freshly obtained or may have been stored under appropriate

temperatures and conditions known in the art. Methods for extracting nucleic acids from a fecal sample are also well known in the art. The extracted nucleic acids may or may not be amplified prior to being used as an input for profiling the relative abundances of bacterial taxa, depending upon the type and sensitivity of the downstream method. When amplification is desired, nucleic acids may be amplified via polymerase chain reaction (PCR). Methods for performing PCR are well known in the art. Selection of nucleic acids or regions of nucleic acids to amplify are discussed above. The nucleic acids comprising the nucleic acid sample may also be fluorescently or chemically labeled, fragmented, or otherwise modified prior to sequencing or hybridization to an array as is routinely performed in the art.

In some embodiments, nucleic acids are amplified using primers that are compatible with use in, e.g., Illumina's reversible terminator method, Roche's pyrosequencing method (454), Life Technologies 's sequencing by ligation (the SOLiD platform) or Life

Technologies's Ion Torrent platform. Examples of such methods are described in the following references: Margulies et al (Nature 2005 437: 376- 80); Ronaghi et al (Analytical Biochemistry 1996 242: 84-9); Shendure et al (Science 2005 309: 1728-32); Imelfort et al (Brief Bioinform. 2009 10:609-18); Fox et al (Methods Mol Biol. 2009;553:79-108);

Appleby et al (Methods Mol Biol. 2009;513: 19-39) and Morozova et al (Genomics. 2008 92:255-64), which are incorporated by reference for the general descriptions of the methods and the particular steps of the methods, including all starting products, reagents, and final products for each of the steps.

In another embodiment, the isolated microbial DNA may be sequenced using nanopore sequencing (e.g., as described in Soni et al. Clin Chem 2007 53: 1996-2001, or as described by Oxford Nanopore Technologies). Nanopore sequencing technology is disclosed in U.S. Pat. Nos. 5,795,782, 6,015,714, 6,627,067, 7,238,485 and 7,258,838 and U.S. Pat Appln Nos. 2006003171 and 20090029477.

The isolated microbial fragments may be sequenced directly or, in some embodiments, the isolated microbial fragments may be amplified (e.g., by PCR) to produce amplification products that sequenced. In certain embodiments, amplification products may contain sequences that are compatible with use in, e.g., Illumina's reversible terminator method, Roche's pyrosequencing method (454), Life Technologies' sequencing by ligation (the SOLiD platform) or Life Technologies' Ion Torrent platform, as described above.

In certain embodiments, the sample sequenced may comprise a pool of nucleic acids from a plurality of samples, wherein the nucleic acids in the sample have a molecular barcode to indicate their source. In some embodiments the nucleic acids being analyzed may be derived from a single source (e.g., from different sites or a time course in a single subject), whereas in other embodiments, the nucleic acid sample may be a pool of nucleic acids extracted from a plurality of different sources (e.g., a pool of nucleic acids from different subjects), where by "plurality" is meant two or more. Molecular barcodes may allow the sequences from different sources to be distinguished after they are analyzed.

In some embodiments, gut microbiota samples are obtained from a subject (e.g., a healthy subject or a not healthy subject (e.g., a patient or a subject in need of treatment according to the technology provided herein) at any suitable interval of time, varying from minutes to hours apart, days to weeks apart, or even weeks to months apart. Gut microbiota samples may be obtained multiple times a day, week, month or year. The duration of sampling can also vary. For example, the duration of sampling may be for about a month, about 6 months, about 1 year, about 2 years, about 3 years, about 4 years, about 5 years, about 6 years, about 7 years, about 8 years, about 9 years, about 10 years, about 11 years, about 12 years, about 13 years, about 14 years, about 15 years, about 16 years, about 17 years, about 18 years, about 19 years, about 20 years, about 30 years, or more.

In some embodiments, a metabolomics screen is performed on a sample from a subject identify, quantify, etc. various metabolite present. In some embodiments, one or more key metabolites are assayed (e.g., metabolites that contribute to sepsis, etc.). In some embodiments, the components of a composition for the treatment of a subject (e.g., the quantity and identity of the Prevotellaceae, Rikenellaceae, Porphyromonadaceae,

Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Prevotella,

Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus bacteria, the quantity and identity of non-bacterial components (e.g., metabolic pathway enzymes, metabolites, etc.)) is determined based on the results of testing (e.g., for metabolites, for enzymatic activity, for microbiota, for sepsis, combinations thereof, etc.).

In some embodiments, a subject or sample therefrom is tested for sepsis (e.g., before and/or after treatment. In some embodiments, more than one test is preformed to diagnose sepsis. In some embodiments, a blood sample is obtained from a subject and tested for evidence of infection (e.g., viral infection, bacterial infection, etc.), clotting problems, abnormal liver or kidney function, impaired oxygen availability, electrolyte imbalances, etc. In some embodiments, blood samples are obtained from two or more distinct sites on a subject and tested for evidence of infection (e.g., viral infection, bacterial infection, etc.), clotting problems, abnormal liver or kidney function, impaired oxygen availability, electrolyte imbalances, etc. In some embodiments, testing for sepsis comprises testing urine (e.g., for signs of bacteria). In some embodiments, testing for sepsis comprises testing wound secretions. In some embodiments, testing for sepsis comprises testing respiratory secretions. In some embodiments, the location of an infection is identified by one or more of X-ray (e.g., of the lungs), Computerized tomography (e.g., to identify infection of, for example, the appendix, pancreas, bowels, etc.), ultrasound, etc.

In some embodiments, subjects identified as being at increased risk of sepsis, subjects identified as suffering from sepsis, and/or subject having gut microbiota that does not promote immune response (e.g., to sepsis), may be administered compositions described herein.

In some embodiments, a subject is treated with a composition comprising

Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Lachnospiraceae, Ruminococcaceae, and/or Bacteroidaceae family bacteria; Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus genera bacteria; and/or bacterial species or strains selected from one or more of Tables 1-6; based on the outcome of a test (e.g., metabolomics testing, microbiomic testing, etc.). Accordingly, in some embodiments, a subject is tested and then treated based on the test results. In some embodiments, a subject is treated and then tested to assess the efficacy of the treatment. In some embodiments, a subsequent treatment is adjusted based on a test result, e.g., the dosage amount, dosage schedule, composition administered, etc. is changed. In some embodiments, a patient is tested, treated, and then tested again to monitor the response to therapy and/or to change the therapy. In some embodiments, cycles of testing and treatment may occur without limitation to the pattern of testing and treating (e.g., test/treat, treat/test, test/treat/test, treat/test/treat, test/treat/test/treat, test/treat/test/treat/test, test/treat/test/test/treat/treat/treat/test, test/treat/treat/test/treat/treat, etc.), the periodicity, or the duration of the interval between each testing and treatment phase.

Although some embodiments herein are described in connection with the treatment or prevention of sepsis, embodiments herein are no limited to as much. Some embodiments herein include the treatment of humans of any suitable age (e.g., infant, child adolescent, adult, etc.) in order to stabilize the subject's microbiota and/or to promote healthy immune system in the subject. EXPERIMENTAL

Example 1

FMT-mediated protection against sepsis

Experiments were conducted during development of embodiments herein we that mimic the development of sepsis in the context of surgical intervention by utilizing a mouse model that involves pre-operative food deprivation, administration of a broad spectrum antibiotic, surgical injury, and introduction of a community of pathogens isolated from a septic critically ill patient, which results in an 80% mortality rate by postoperative day 3 (POD3) (Alverdy 2000; Laughlin 2000; Zaborin AAC 2014; incorporated by reference in their entireties) (Fig. 1A-B). Using this model, experiments demonstrate that FMT leads to a striking increase in survival, and this protective effect is mediated by an increase in the abundance of specific taxa that are also associated with the replenishment of gut microbial diversity and the resulting restoration of immune homeostasis. Experiments confirm the protective effects of FMT and the relative proportions of the relevant taxa in the proposed formulation in a second model where dissemination to peripheral organs has already taken place, demonstrating that restoring the microbiome is hugely protective even when the systemic response to sepsis is already established.

Materials and Methods

Bacterial preparation

The pathogen community (PC) contained four members including Candida albicans, Enter ococcus faecalis, multi-drug resistant Klebsiella oxytoca, and multi-drug resistant Serratia marcescens and was isolated from the stool of a critically ill patient who was exposed to numerous antibiotics during the course of their sepsis (Zaborin MBio 2014; Zaborin AAC, 2014; incorporated by reference in their entireties). The individual pathogens were plated on TSB agar from their individual frozen stocks and grown overnight at 37^°C.

Colonies were suspended in liquid TSB medium, subgrown for 1 hour and then adjusted to an OD600 of 0.2. All four microbe species were then combined together in equal volumes. Bacterial suspension was centrifuged at 6,000 rpm for 10 min, the excess TSB removed and the remaining pellet was re-suspended in the same volume of 10% glycerol. The resulting microbial suspension was used for cecal injections, as described below. Mice

Seven to nine week old C57BL/6 mice (Charles River) weighing 18-22 g were used for experiments. Mice used in comparative studies were the progeny of IRF3^+/" mice resulting in IRF3^+/+ and IRF3^_/" littermates.

Mouse model of lethal gut-derived sepsis

All experiments were approved by the Institutional Animal Care and Use Committee at the University of Chicago (IACUC protocol 71744). Mice were routinely fed tap water and Harland Teklad feed (Madison, WI) under 12-h light/dark cycles and were allowed to acclimate for at least 48 hr before surgery. In order to mimic pre-operative surgical conditions, mice were exposed to food restriction and prophylactic antibiotics.

Approximately 16 hours prior to surgery, mice were moved to wire floor cages to prevent coprophagy. Food was removed, and mice were allowed only tap water ad libitum. Each mouse received an intramuscular Cefoxitin injection 30 minutes prior to the incision at a concentration of 25 mg/kg into the left thigh. Mice were then subjected to a midline laparotomy, 30% hepatectomy of the left lateral lobe of the liver and a direct cecal inoculation of PC (200 of PC suspension prepared as described above). On the day of surgery, a fresh fecal microbiota transplant (FMTii_ve) was prepared by harvesting the cecal contents of normal healthy mice. Cecal contents were then suspended in sterile 10% glycerol at a concentration of 50 mg/mL. An aliquot of the cecal contents was autoclaved and resuspended in sterile 10% glycerol to a final concentration of 50 mg/ml to create the autoclaved control (FMT_autocl). On postoperative days one and two (POD1 and POD2), randomly selected group of mice were given either 1 ml of fecal microbiota transplant

(FMTii_ve) or FMTautocl via rectal enema using 10 Fr pediatric red rubber catheter. Chow was restored to the cages on postoperative day two, approximately 45 hours after the operation. Mice were examined every 6 hours and scored for illness based on a sepsis scoring system. Score 1- alert, responsive, and normal activity; Score 2- poor grooming or decreased activity; Score 3- decreased activity but moves when stimulated, ruffled fur or hunched posture increased respiratory rate; Score 4- fails to move when touched difficulty breathing or irregular respiratory partem; Score 5- dead. All mice that were moribund were immediately sacrificed and considered to be a mortality. 16SrRNA analysis

Microbial DNA was extracted from cecal luminal contents and cecal tissues using Bacteremia DNA Isolation Kit (BiOstic, 12240-50), and the 16S rRNA analysis was performed at Argonne National Laboratories (Lemont, IL). Samples were analyzed by 16S rRNA V4 iTAG amplicon sequencing analysis. Paired end reads were quality trimmed and processed for OTU (operational taxonomic unit) clustering using UP ARSE pipeline (Edgar, 2013; incorporated by reference in its entirety), set at 0.97% identity cutoff. Taxonomic status was assigned to the high quality (<\% incorrect bases) candidate OTUs using the

"parallel_assign_taxonomy_rdp.py" script of QIIME software (Caporaso et al, 2010;

incorporated by reference in its entirety). Multiple sequence alignment and phylogenetic reconstruction was performed using PyNast and FastTree (Caporaso et al, 2010;

incorporated by reference in its entirety). OTU matrix was processed to remove OTUs containing less than 5 reads in order to reduce the PCR and sequencing based bias; then the OTU table was rarified to the minimum numbers of reads present in the smallest library. The oligotyping pipeline (Eren et al 2013; incorporated by reference in its entirety) was used to identify the sub-OTU level differences in the introduced pathogen strains of Klebsiella oxytoca ICUl-2, Serratia marcescens ICUl-2 and Enterococcus faecalis ICUl-2. Full length 16S rRNA gene sequence were extracted from genome sequences of K. oxytoca, S.

marcescens ICUI-2 and E. faecalis genomes using Blastn. Megablast (minimum identity cut- off=100%) was used to confirm the strain identification between full length 16S rRNA gene sequences and oligotype representative sequences.

Sequencing of bacterial pathogen genomes

For isolation of DNA, strains of the original stock consisting of Klebsiella oxytoca, Serratia marcescens and Enterococcus faecalis were collected from exponential phase during growth in liquid TSB. Sequence libraries were generated using Nextera XT protocol according to manufacturer's instructions (Illumina). Libraries were sequenced by whole- genome shotgun sequencing using the Illumina HiSeq system at Argonne National

Laboratory (Lemont, Chicago). Approximately 150-fold coverage of each genome was generated. These reads were quality trimmed and assembled using nesoni

(github.com/Victorian-Bioinform and Velvet (version 1.2.10), respectively. Draft genome assemblies (minimum contig length = 300bp) have been submitted to NCBI. Accession numbers in NCBI are LQAM00000000 Enter ococcus, LQAL00000000 Klebsiella, and LQAK00000000 Serratia in the BioProject PRJNA307050.

Evaluation of systemic infection

Blood, spleen and liver were assessed for colonization with PC species using selective plates: for C. albicans, CHROMagar Candida plates (BBL); ΐοχ K. oxytoca, MacConkey supplemented with ciprofloxacin, 10 μg/ml; for S. marcescens, MacConkey supplemented with imipenem, 1 μg/ml; and for E. faecalis , Enterococcal agar plates (BBL). The specific Gram (-) negative plates supplemented with antibiotics were chosen for K. oxytoca ICUl-2 and S. marcescens ICUl-2 selection based on their antibiotic resistance, and the selectivity was verified by plating intestinal content of untreated mice with no growth on the plates. CHROMagar Candida plates select for C. albicans seen as blue colonies. Enterococcal agar plates select for Enterococcus seen as black colonies. Blood was collected via direct cardia puncture, mixed with glycerol to a final concentration 10%, and stored at -80°C. 10 mg of liver and spleen were homogenized in 500 μΐ of 10% glycerol and stored in -80°C. Each sample was serially diluted, and 50 μΐ of each dilutions were plated on above mentioned four selective plates. All the plates were incubated in 37°C incubator for 24 hours. CFU counts were normalized to tissue weight for spleen and liver and volume for blood. Cecal Content Lysates Preparation

Healthy mice were sacrificed according to approved IACUC protocols. Ceca were resected and cecal contents were isolated, centrifuged for 5 min at 6000 rpm and the supernatant was discarded. The pellet was resuspended in 500 μΐ PBS and 100 μΐ of zirconia beads were added. The samples were then vortexed horizontally using Ambion Vortex Adapter for Vortex-Genie 2 (Life Technologies) at maximum speed for 10 minutes. The suspension was then centrifuged at 10,000 rpm for lmin and the supernatant was removed and filtered using 0.22μιη filter and stored at -80^°C for later use. Prior to use, protein concentration of the lysates was measured with BCA Protein Assay Kit (Pierce™) and normalized to 25 μg/ml.

Splenic Dendritic Cell Cytokine Production

Splenic dendritic cells (DCs) were isolated as described previously (Devkota et al, Nature 2012; incorporated by reference in its entirety). Spleens were harvested from 6-8 week old C57BL/6 mice, mashed them into a single cell suspension, lysed red blood cells (R&D Systems), and positively selected for CD1 lc⁺ cells to a purity of up to 90% using MACS beads according to manufacturer protocol (Miltenyl Biotec). 10⁶ DCs were cultured in 24 well plates with 2ng/mL of TGF-β and O. lnM retinoic acid. 25 μg of cecal content lysate was added and cells were cultured for 24h at 37C, after which supernatant cytokine levels were measured by ELISA. Π β (Duoset, R&D Systems), and IL-6, IL-10, IL-12p40, IL- 12p70, TNFa (OptiEIA, BD Biosciences) levels were measured by sandwich ELISA according to manufacturer protocols. Mouse model of lethal sepsis due to intraperitoneal (IP) injection of the PC

In order to determine if the rescue effect of FMT functions at the level systemic infection, mice were directly injected with the PC into the peritoneum (IP). In the IP model, mice were not subjected to starvation, antibiotic treatment or hepatectomy. The suspension was prepared using the identical strains (S. marcescens, C. albicans, K. oxytoca, and E. faecalis) and methods above. The PC was plated on TSB plates and grew overnight at 37^°C.

Few colonies from each species were dissolved in 10% glycerol, adjusted to 0.2 OD and 50 μΐ of each prepared solution were mixed in 1 ml of 10% glycerol that was then injected intraperitoneally. The optimal and lethal dose of PC was determined by preliminary experiments. FMTii_ve and FMT_aut0ci were prepared as described above. Each mouse received two doses of either FMTii_ve or FMT_aut0ci, immediately after IP injection and 12 hours later. Mice were examined every 6 hours and scored for illness based upon sepsis scoring system. Score 1- alert, responsive, and normal activity; Score 2- poor grooming or decreased activity; Score 3- decreased activity but moves when stimulated, ruffled fur or hunched posture increased respiratory rate; Score 4- fails to move when touched difficulty breathing or irregular respiratory partem; Score 5- dead. All mice that were moribund were immediately sacrificed and considered to be a mortality.

Mouse whole-genome transcription analysis

The cecum, left lobe of the liver, and spleen were isolated from mice and stored in

RNAprotect cell reagent (Qiagen). RNA extraction was performed using the RNeasy Plus Mini Kit (Qiagen). Whole-genome transcriptional profiling was performed using Illumina MouseRef-8 v2 arrays (Illumina) by Functional Genomics Facility of the University of Chicago. Data were quantile normalized using Genome Studio 201 lvl. Selection of differentially expressed genes (DEGs) was based on pair-wise comparisons of control (untreated) and treated groups of mice using Significant Analysis of Microarrays (SAM) with False Discovery Ratio <\% and minimal fold changes +/-2.0 (Tusher VG, Tibshirani R, Chu G, PNAS, 2001). Pathways and functions associated with selected sets of DEGs were identified using Ingenuity Pathway Analysis software (https://analysis.ingenuity.com/) using default settings. Presented data are based on the functions "Canonical Pathways" and "network analysis". Cluster analysis was performed with JMP 11 software (SAS, Cary, NC) using Ward method as linkage metric and Euclidian distances as similarity metric. Quantitative-PCR

RNA was isolated from cecum, liver, and spleen as before and converted to cDNA using reverse transcriptase (Promega). The qPCR primers targeting the MASV, IRF3, USP21, TRAF6 genes were custom ordered from Integrated DNA Technologies. qPCR was performed with SYBR Advantage qPCR Premix (Clontech) on a LightCycler 480 (Roche). Expression of all genes was normalized to that of the housekeeping gene GAPDH.

Statistical analysis

Statistical analysis was performed using SigmaPlot software. Student's t test was used when analyzing the differences between two means, whereas analysis of variance (ANOVA) with Bonferroni correction was used when more than two means were compared.

Nonparametric Wilcoxon-Mann-Whitney test was used to determine the statistical significance of culture analyses. Kaplan-Meier survival curves were analyzed using SPSS software. Significance was determined as a P value <0.05. Results

FMT rescues mice from gut-derived sepsis

It has been demonstrated that all elements of the process of major surgery are required for the development of lethal gut-derived sepsis in a mouse model that includes pre-operative food deprivations (i.e. H₂0 only), pre-operative antibiotic administration (cefoxitin), surgical injury (30% hepatectomy), and colonization by intestinal pathogens (Fig. 1A; Alverdy 2000; Laughlin 2000; Zaborin AAC 2014; incorporated by reference in their entireties). The intestinal pathogen community used in this study (denoted PC hereafter) was taken from the cecum of a patient that had severe sepsis and is comprised of three species of bacteria (Enter ococcus faecalis, multi-drug resistant Klebsiella oxytoca, and multi-drug resistant Serratia marcescens) and one species of yeast {Candida albicans) that were injected into the cecum of mice concurrently with surgical injury (Fig. 1A). Consistent with previous reports (Zaborin AAC 2014; incorporated by reference in its entirety), this model of gut- derived sepsis resulted in an 80% mortality rate by postoperative day 3 (POD3) (Fig. IB). To investigate whether FMT was protective in this model, reiterative experiments were performed by randomly assigning septic mice at POD1 to two groups. One group was administered a rectal enema of a live fecal microbial transplant (FMTii_ve) consisting of pooled cecal contents harvested from healthy mice. The control group was administered a rectal enema of the same material, only the cecal contents were autoclaved (FMT_aut0d) to demonstrate the need for live bacterial transplantation (Fig. 1A). The enema administration was repeated on POD2 in both groups. Remarkably, administration of FMTii_ve led to approximately a 60% increase in survival (from 20% to 80%), whereas no protection was observed with FMT_aut0ci (Fig. IB). FMT prevents the progression to a pathobiome and restores microbial homeostasis

To investigate the impact of FMT on the gut microbiota of septic mice, 16S rRNA analysis of cecal luminal bacteria was performed in a temporal manner. Analysis of microbial diversity demonstrates a significant decrease in diversity in the cecal microbiota of mice at POD1 before FMTii_ve or FMT_aut0ci were administered (Fig. 2A). Comparison of diversity indices between FMTii_ve and FMT_aut0ci groups demonstrates that the microbiota of FMTii_ve treated mice had higher diversity at both POD2 and POD3. By POD7, the diversity of the FMTii_ve group was similar to that of untreated mice. Given that most of FMT_autoci group mice showed died before POD7, results do not include samples from POD7 among FMT_aut₀ci mice.

Analysis of the relative abundance of taxa at the genus (Fig. 2B) level demonstrates that mice subjected to pre-operative starvation and antibiotic treatment (i.e. POD0) had significant alterations to microbiome composition, with a notable increase of Allobaculum and decrease of Lactobacillus in the cecum (Fig. 2B). Twenty-four hours after hepatectomy and intestinal inoculation of the pathogen community (POD1), there was domination of Enter ococcus and Allobaculum (Fig. 2B). Treatment with FMTii_ve significantly shifted microbiota composition on POD2 and POD3. Compared to mice that continue to progress to sepsis (i.e. FMT_aut0Ci), mice treated with FMTii_ve have a significant decrease in the relative abundance of Serratia followed by its complete elimination by POD7, with significant reappearance of microbiota representative of the preoperative state (Fig. 2B). Importantly, the bacterial families S24-7 (Prevotella-like), Rikenellaceae, Prevotellaceae, Porphyromonadaceae, Lactobacillaceae, and Bacteroidaceae are significantly associated with immune system activation and stable immune-inflammation response. In the field several species of the family Ruminococcaceae have been miss-identified and belong to the family Lachnospiraceae. Therefore, in some embodiments, the bacteria of the family

Lachnospiraceae are withint he scope herein.

To precisely determine whether the injected PC members successfully colonized the cecum of septic mice, we utilized the oligotyping pipeline (Eren et al., 2013; incorporated by reference in its entirety) to compare the sequences of the cecal luminal microbiota to that of the inoculating pathogens on a sub-OTU level. The presence of S. marcescens (Fig. 2C), E. faecalis (Fig. 2D), and K. oxytoca (Fig. 2E) with 100% similarity to the inoculating strain (Oligotype 1) was seen from POD1-POD3 in both FMT_aut0d and FMTii_ve mice. Importantly, in FMTiive treated mice, there was a marked reduction in Oligotype 1 of all three bacterial pathogens over time, and by POD7 the inoculating pathogens were completely absent from the guts of these mice.

In order to determine the impact of these differences of microbiota in terms of inflammatory potential, we isolated cecal contents from untreated, FMTii_ve -POD2 and FMTautod -POD2 mice, and used these to stimulate splenic dendritic cells (DCs) (Fig. 2F) and bone marrow-derived DCs in vitro. The cecal contents from FMT_aut0ci mice drove the production of IL-Ιβ, IL6, IL10, and IL12p70 by DCs (Fig. 2F). Meanwhile the cecal contents of FMTii_ve led to cytokine production that was similar to cecal contents from untreated mice. Overall, these results demonstrate that treatment of septic mice with FMTii_ve leads to a restoration of microbial diversity and a reduction in pathogens, and the microbiota of septic mice treated with FMTii_ve shows a significant reduction in immune-stimulatory capacity. FMT protects against mortality in mice following systemic inoculation of the PC

The results obtained in the gut-derived sepsis model led to the question whether FMTiive is leading to the restoration of systemic homeostasis by merely blocking the dissemination of pathogens from the gut, or whether it was also actuating a host response that drives pathogen clearance. To dissociate these two possibilities, a second lethal model of sepsis was developed where the PC is injected directly into the peritoneal cavity (i.p.) of mice to presumably cause immediate systemic dissemination of pathogens (IP model). As before, mice were treated with either FMTii_ve or FMT_aut0ci via enema on two consecutive days; on the day of pathogen inoculation and on the following day; however, there was no fasting, surgery, or application of antibiotics (Fig. 3A). Results demonstrated that both the appearance of sepsis and mortality were significantly attenuated by the administration of FMTii_ve (n=15/group, p<0.001) (Fig. 3B) and significant clearance of all the PC members was observed from all of the three organs, except for C. albicans in blood (Fig. 3C-F). Thus, even in a situation where pathogens are inoculated systemically, the administration of FMTii_ve into the gut can rescue mice from otherwise lethal sepsis and drive the systemic clearance of pathogens.

In the gut-derived sepsis model, the most differentially regulated pathways associated with the progression of sepsis are the RIG-I and IRF3 signaling pathways. To determine if these pathways were also significantly changed in the IP model, and if FMTii_ve mediated protection in this model was associated with the reversion of these pathways to homeostasis, we examined the expression patterns of key RIG-I / IRF pathway genes. Expression patterns of MAVS, IRF3, Interferon stimulated gene USP21, and TRAF6 were analyzed by quantitative RT-PCR in samples obtained at sacrifice from the cecum, liver, and spleen of untreated mice, FMTautoci on POD2, and FMTii_ve on POD2. For comparison, the data were compared with microarray data of the gene expression in the cecum, liver and spleen in the hepatectomy model. In concordance with results from the gut-derived sepsis model, similar to previous model of lethal gut-derived sepsis, results from the IP model demonstrated that the expression of MAVS, IRF3, and USP21 was down-regulated in the FMT_aut0ci group but maintained or reverted to control normal levels (i.e. compared comparable to untreated mice) in the FMTli_ve group. The expression of TRAF6 was up-regulated in the FMT_aut0ci group and maintained at control levels or returned to normal levels in the FMTii_ve group (Fig. 3G-J). Based on the fact that we see in both models the dramatic modulation of these pathways with the progression of sepsis, and that FMT-mediated protection coincides with a reversion of these genes to baseline levels, we hypothesized that these pathways play a central role in FMT-mediated protection against sepsis. Taken together these results confirm that the FMTii_ve can modulate the systemic immune response in association with clearance of microbial pathogens that have disseminated systemically to organs.

Example 2

Immunomodulatory strain selection

Based on comparative metagenome analysis described above, the taxonomic analysis of raw and assembled metagenomic reads indicates that bacteria from the families

Prevotellaceae, Rikenellaceae plus Lachnospiraceae, Porphyromonadaceae, Lactobacillaceae, and Bacteroidaceae play roles in supporting the immune response of a subject, e.g. under conditions of a bacterial infection that have the potential to develop into a sepsis response. Therefore, strains were selected from these bacterial families, based on strain characteristics, that will find use in biotherapeutic formulations for modulating the immune response in a subject. The major fermentation products produced by strains isolated from the human gut 5 microbiome were used to make an initial strain selection among the 773 strains currently present in the Virtual Metabolic Human database (vmh.uni.lu/#microbes/search). Genome annotation and in silico modeling was subsequently performed to confirm the publicly available information for these fermentation products. The genome annotation platform, RAST, was used to confirm the presence of the key functionalities in the strains resulting 0 from the analysis shown in FIG. 2B of microbiota representative of the preoperative state.

These bacterial families are significantly associated with immune system activation and stable immune-inflammation response.

5 Table 1: Overview of strains belonging to the family Prevotellaceae, represented by strains belonging to the genera Prevotella and Paraprevotella, to be included in a live biotherapeutic formulation to modulate an immune response in a subject.

Bacteroides Bacteroidaceae Iso-But + Iso-Val Human feces coprophilus DSM

18228

Bacteroides dorei Bacteroidaceae Iso-But + Iso-Val Human feces

DSM 17855

Bacteroides eggerthii Bacteroidaceae Iso-But + + Human feces; also produces

DSM 20697 phenylacetate

Bacteroides faecis Bacteroidaceae + + Human feces

DSM 24798

Bacteroides Bacteroidaceae Iso-But + Iso-Val Human feces finegoldii DSM

17565

Bacteroides fluxus Bacteroidaceae + + Human feces

DSM 22534

Bacteroides Bacteroidaceae Iso-But + + Iso-Val Human feces intestinalis DSM

17393

Bacteroides Bacteroidaceae + + Human feces oleiciplenus DSM

22535T

Bacteroides ovatus Bacteroidaceae Iso-But + Iso-Val Human feces; also produces

DSM 1896 phenylacetate

Bacteroides Bacteroidaceae + Human feces pectinophilus ATCC

43243

Bacteroides plebeius Bacteroidaceae Iso-But + Iso-Val Human feces

DSM 17135

Bacteroides stercoris Bacteroidaceae Iso-But + + Iso-Val Human feces; also produces

DSM 19555 phenylacetate

Bacteroides Bacteroidaceae Iso-But + + Human feces; also produces thetaiotaomicron phenylacetate

DSM 2079

Bacteroides Bacteroidaceae + + Human feces timonensis DSM

26083

Bacteroides Bacteroidaceae Iso-But Iso-Val Human feces; also produces uniformis DSM 6597 phenylacetate

Bacteroides vulgatus Bacteroidaceae Iso-But + Iso-Val Human feces

DSM 1447

Bacteroides Bacteroidaceae Iso-But + + Iso-Val Human faces xylanisolvens DSM

18836

Table 2: Overview of strains belonging to the family Bacteroidaceae, represented by strains belonging to the genera Parabacteroides and Bacteroides, to be included in a live biotherapeutic formulation to modulate an immune response in a subject.

5

ATCC 25644

Lactobacillus ultunensis Lactobacillaceae + Human stomach mucosa DSM 16047

Table 3: Overview of strains belonging to the family Lactobacillaceae, represented by strains belonging to the genus Lactobacillus, to be included in a live biotherapeutic formulation to modulate an immune response in a subject.

5

Table 4: Overview of strains belonging to the family Ruminococcaceae, represented by strains belonging to the genus Ruminococcus, to be included in a live biotherapeutic formulation to modulate an immune response in a subject.

0 The Rikenellaceae is a recently established family of bacteria and comprises the genus

Rikenella and Alistipes. Rikenella is monospecific and contains the species R. microfusus, which was previously classified as Bacteroides microfusus. The genome sequence of a representative strain of this species, such as R. microfusus DSM 15922, is currently not available. Strains belonging to the genus Alistipes to be considered in a life biotherapeutic 5 formulation to modulate the immune response in a subject, are listed in Table 5.

Table 5: Overview of strains belonging to the family Rikenellaceae, represented by strains belonging to the genus Alistipes, to be included in a life biotherapeutic formulation to modulate an immune response in a subject.

0 Several species of the family Ruminococcaceae have been miss-identified and belong to the family Lachnospiraceae. Therefore, the selection of strains should also include this family. Examples are presented in Table 6.

5 Table 6: Overview of strains belonging to the family Lachnospiraceae, represented by strains belonging to the genera Anaerostipes, Blautia, Clostridium, Coprococcus, Dorea, Eubacterium, Marvinbryantia and Roseburia, to be included in a life biotherapeutic formulation to modulate an immune response in a subject. ¹ The propionate synthesis observed for members of the Lachnospiraceae is often the result of promiscuous enzyme specificities and in limited quantities.

REFERENCES

The following references, some of which are cited above, are herein incorporated by reference in their entireties. Alverdy, J., Holbrook, C, Rocha, F., Seiden, L., Wu, R.L., Musch, M., Chang, E., Ohman, D., and Suh, S. (2000). Gut-derived sepsis occurs when the right pathogen with the right virulence genes meets the right host: evidence for in vivo virulence expression in Pseudomonas aeruginosa. Ann. Surg. 232, 480-489.

Angus, D.C., and van der Poll, T. (2013). Severe sepsis and septic shock. N. Engl. J. Med.

369, 840-851.

Angus, D.C., Linde-Zwirble, W.T., Lidicker, J., Clermont, G, Carcillo, J., and Pinsky, M.R.

(2001). Epidemiology of severe sepsis in the United States: analysis of incidence, outcome, and associated costs of care. Crit. Care Med. 29, 1303-1310.

Cammarota, G, Ianiro, G., and Gasbarrini, A. (2014). Fecal microbiota transplantation for the treatment of Clostridium difficile infection: a systematic review. J. Clin.

Gastroenterol. 48, 693-702.

Caporaso, J.G, Bittinger, K., Bushman, F.D., DeSantis, T.Z., Andersen, G.L., and Knight, R.

(2010). PyNAST: a flexible tool for aligning sequences to a template alignment.

Bioinformatics 26, 266-267.

Colman, R.J., and Rubin, D.T. (2014). Fecal microbiota transplantation as therapy for

inflammatory bowel disease: a systematic review and meta-analysis. J Crohns Colitis 8, 1569-1581.

Devkota, S., Wang, Y., Musch, M.W., Leone, V., Fehlner-Peach, H., Nadimpalli, A.,

Antonopoulos, D.A., Jabri, B., and Chang, E.B. (2012). Dietary-fat-induced taurocholic acid promotes pathobiont expansion and colitis in 1110-/- mice. Nature 487,

104-108.

Edgar, R.C. (2013). UP ARSE: highly accurate OTU sequences from microbial amplicon

reads. Nat. Methods 10, 996-998.

Eiseman, B., Silen, W., Bascom, G.S., and Kauvar, A.J. (1958). Fecal enema as an adjunct in the treatment of pseudomembranous enterocolitis. Surgery 44, 854-859. Eren, A.M., Maignien, L., Sul, W.J., Murphy, L.G., Grim, S.L., Morrison, H.G, and Sogin, M.L. (2013). Oligotyping: Differentiating between closely related microbial taxa using 16S rRNA gene data. Methods Ecol Evol 4.

Honda, K., and Taniguchi, T. (2006). IRFs: master regulators of signalling by Toll-like

receptors and cytosolic pattern-recognition receptors. Nat. Rev. Immunol. 6, 644-658.

Iwasaki, A., and Medzhitov, R. (2010). Regulation of adaptive immunity by the innate

immune system. Science 327, 291-295.

Lagu, T., Rothberg, M.B., Shieh, M.-S., Pekow, P.S., Steingrub, J.S., and Lindenauer, P.K.

(2012). Hospitalizations, costs, and outcomes of severe sepsis in the United States 2003 to 2007. Crit. Care Med. 40, 754-761.

Laughlin, R.S., Musch, M.W., Hollbrook, C.J., Rocha, F.M., Chang, E.B., and Alverdy, J.C.

(2000). The key role of Pseudomonas aeruginosa PA-I lectin on experimental gut- derived sepsis. Ann. Surg. 232, 133-142.

van Nood, E., Vrieze, A., Nieuwdorp, M., Fuentes, S., Zoetendal, E.G., de Vos, W.M., Visser, C.E., Kuijper, E.J., Bartelsman, J.F.W.M., Tijssen, J.G.P., et al. (2013). Duodenal infusion of donor feces for recurrent Clostridium difficile. N. Engl. J. Med. 368, 407- 415.

Pigneur, B., and Sokol, H. (2016). Fecal microbiota transplantation in inflammatory bowel disease: the quest for the holy grail. Mucosal Immunol.

Smits, L.P., Bouter, K.E.C., de Vos, W.M., Borody, T.J., and Nieuwdorp, M. (2013).

Therapeutic potential of fecal microbiota transplantation. Gastroenterology 145, 946-

953.

Sorensen, T.I., Nielsen, G.G, Andersen, P.K., and Teasdale, T.W. (1988). Genetic and

environmental influences on premature death in adult adoptees. N. Engl. J. Med. 318, 727-732.

Tusher, V.G, Tibshirani, R., and Chu, G. (2001). Significance analysis of microarrays

applied to the ionizing radiation response. Proc. Natl. Acad. Sci. U.S.A. 98, 5116- 5121.

Vrieze, A., Van Nood, E., Holleman, F., Salojarvi, I, Kootte, R.S., Bartelsman, J.F.W.M., Dallinga-Thie, G.M., Ackermans, M.T., Serlie, M.J., Oozeer, R, et al. (2012).

Transfer of intestinal microbiota from lean donors increases insulin sensitivity in individuals with metabolic syndrome. Gastroenterology 143, 913-916.e7.

Zaborin, A., Defazio, J.R., Kade, M., Kaiser, B.L.D., Belogortseva, N., Camp, D.G, Smith, R.D., Adkins, J.N., Kim, S.M., Alverdy, A., et al. (2014a). Phosphate-containing polyethylene glycol polymers prevent lethal sepsis by multidrug-resistant pathogens. Antimicrob. Agents Chemother. 58, 966-977.

Zaborin, A., Smith, D., Garfield, K., Quensen, I, Shakhsheer, B., Kade, M., Tirrell, M., Tiedje, J., Gilbert, J.A., Zaborina, O., et al. (2014b). Membership and behavior of ultra-low-diversity pathogen communities present in the gut of humans during prolonged critical illness. MBio 5, e01361-01314.

Zhang, F., Luo, W., Shi, Y., Fan, Z., and Ji, G. (2012). Should we standardize the 1,700-year- old fecal microbiota transplantation? Am. J. Gastroenterol. 107, 1755; author reply p.1755-1756.

Claims

1. A method of modulating an immune response in a subject, the method comprising administering to the subject a composition comprising bacteria selected from: one or more of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae,

Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae; one or more of the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus; and/or bacteria of the species and/or strains listed in Table 1, Table 2, Table 3, Table 4, Table 5, and/or Table 6.

2. The method of claim 1, wherein the modulation of the immune response comprises preventing infections.

3. The method of claim 2, wherein the modulation of the immune response comprises preventing sepsis.

4. The method of claim 1, wherein the modulation of the immune response comprises treating an infection.

5. The method of claim 4, wherein the modulation of the immune response comprises treating sepsis.

6. The method of claim 2, wherein the subject is at risk of developing sepsis.

7. The method of claim 5, wherein the subject suffers sepsis.

8. The method of claim 7, wherein the subject suffers from severe sepsis or septic shock.

9. The method of one of claims 1-8, wherein the composition comprises bacteria from 5 or more different taxa.

10. The method of claim one of claims 1-9, wherein the composition comprises at least 10⁴ colony forming units (CFU) of bacteria.

11. The method of one or claims 1-10, wherein the subject has abnormal gut microbiota.

12. The method of one or claims 1-11, wherein the subject is a human or an animal.

13. The method of one or claims 1-12, wherein the composition is administered orally.

14. The method of one or claims 1-12, wherein the composition is administered rectally.

15. The method of one or claims 1-14, further comprising assaying the microbiome and/or metabolome of the subject.

16. The method of claim 15, wherein assaying the microbiome comprises testing the presence, absence, or amount of one or more bacteria in the gut of the subject.

17. The method of claim 16, wherein assaying the microbiome comprises testing the presence, absence, or amount of one or more bacteria from one or more of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae; one or more of the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus; and/or bacteria of the species and/or strains listed in Table 1, Table 2, Table 3, Table 4, Table 5, and/or Table 6.

18. The method of claim 15, wherein assaying the metabolome comprises quantifying amount of one or more metabolites in the gut of the subject.

19. The method of claim 15, wherein the assaying is performed on the subject before and/or after administration of the composition.

20. The method of one or claims 1-19, wherein the composition is co-administered with one or more additional active agents.

21. The method of claim 20, wherein the additional active agent comprises a probiotic component, a prebiotic component, or a small molecule with therapeutic activity.

22. The method of claim 20, wherein the additional active agent comprises a microbe.

23. The method of claim 20, wherein the additional active agent comprises an antibody or antibody fragment.

24. A pharmaceutical composition comprising bacteria from one or more of the families Prevotellaceae, Rikenellaceae, Porphyromonadaceae, Lactobacillaceae, Ruminococcaceae, Lachnospiraceae, and/or Bacteroidaceae; one or more of the genera Prevotella, Paraprevotella, Alistipes, Bacteroides, Lactobacillus, Parabacteroides, and/or Ruminococcus; and/or bacteria of the species and/or strains listed in Table 1 , Table 2, Table 3, Table 4, Table 5, and/or Table 6.

25. The pharmaceutical composition of claim 24, comprising a therapeutically effective amount of bacteria.

26. The pharmaceutical composition of claim 24, wherein a therapeutically effective amount of bacteria is an amount sufficient to treat or prevent sepsis in a subject.

27. The pharmaceutical composition of claim 24, wherein a therapeutically effective amount of bacteria is an amount sufficient to activate regulator T cell accumulation in the subject.

28. The pharmaceutical composition of claim 24, wherein the composition comprises at least 10⁴ colony forming units (CFU) of bacteria.

29. The pharmaceutical composition of claim 24, further comprising a probiotic, a prebiotic, or a therapeutically active small molecule.

30. The pharmaceutical composition of claim 24, wherein the bacteria are alive.

31. The pharmaceutical composition of claim 24, wherein the bacteria are sporulated.

32. The pharmaceutical composition of claim 24, formulated for oral administration.

33. The pharmaceutical composition of claim 24, formulated for rectal administration.

34. The pharmaceutical composition of claim 24, wherein the pharmaceutical composition is a nutraceutical or a food.

35. Use of a composition of claim 24 to manufacture a medicament for administration to a subject.

36. Use of a composition of claim 24 to treat or prevent infection in a subject.

37. Use of a composition of claim 24 to treat or prevent sepsis in a subject.