US 20160310546A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2016/0310546A1 Lynch et al. (43) Pub. Date: Oct. 27, 2016

(54) SNUSTS DAGNOSTICS AND Publication Classification TREATMENTS (51) Int. Cl. (71) Applicant: The Regents of the University of A 6LX 35/747 (2006.01) California, Oakland, CA (US) A6IR 9/00 (2006.01) CI2O I/06 (2006.01) (72) Inventors: Susan Lynch, Piedmont, CA (US); A6II 35/744 (2006.01) Andrew Goldberg, Mill Valley, CA (52) U.S. Cl. (US); Steve D. Pletcher, Mill Valley, CPC ...... A61K 35/747 (2013.01); A61K 35/744 CA (US) (2013.01); A61K 9/0043 (2013.01); C12O I/06 (2013.01) (21) Appl. No.: 15/203,428 (22) Filed: Jul. 6, 2016 (57) ABSTRACT Related U.S. Application Data Provided herein are compositions for improving sinus (63) Continuation of application No. 14/394,006, filed on microbiota and treating sinusits. Further provided are meth Oct. 10, 2014, filed as application No. PCT/US2013/ ods of detecting imbalance in the sinus microbiota that can 036297 on Apr. 12, 2013. be indicative of sinusitis, and methods of determining (60) Provisional application No. 61/624,105, filed on Apr. whether an individual has or is at risk of developing sinus 13, 2012. itis, e.g., chronic sinusitis. Patent Application Publication Oct. 27, 2016 Sheet 1 of 8 US 2016/0310546 A1

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SNUSTS DAGNOSTICS AND BRIEF SUMMARY OF THE INVENTION TREATMENTS 0006. Described herein are results from a high-resolution, culture-independent comparative analysis of the sinus CROSS-REFERENCE TO RELATED microbiota of chronic rhinosinusitis (CRS) patients, and APPLICATIONS healthy Subjects without CRS undergoing open nasal or 0001. This application is a continuation of U.S. applica sinus Surgery. Surgical patients typically exhibit severe tion Ser. No. 14/394,006 filed Oct. 10, 2014, which is a S371 disease, and Surgery provides access to affected sinus US National Stage of PCT/US2013/036297 filed Apr. 12, mucosal surfaces that are otherwise inaccessible. The results 2013, which claims priority to U.S. Application No. 61/624, indicate that specific features of the sinus mucosal micro 105 filed Apr. 13, 2012, the disclosures of which are biota are associated with disease state and severity, and incorporated by reference in their entirety. identify both pathogenic and protective species in this niche. 0007 Provided herein are novel probiotics for improving REFERENCE TO SUBMISSION OF A the sinus microbiota in an individual comprising at least one SEQUENCE LISTING AS AN ASCII TEXT FILE bacteria listed in Table X, e.g., a Lactic Acid Bacterial (LAB) species, a Lactobacillus species, or L. Sakei. Such 0002. The Sequence Listing written in file 48536 probiotics can be used to treat an individual with reduced 518C01 US ST25.TXT, created on Jul. 1, 2016, 2,382 bytes, microbial diversity in the nasal sinus, or one that has been machine format IBM-PC, MS-Windows operating system, is diagnosed with acute or chronic sinusitis. In some embodi hereby incorporated by reference. ments, provided is a pharmaceutical composition compris ing at least one bacterial species listed in Table X, and a BACKGROUND OF THE INVENTION pharmaceutically acceptable excipient, e.g., for nasal or 0003. Sinusitis (rhinosinusitis) is one of the most com sinus administration. In some embodiments, the composi mon problems presented to the primary care practitioner in tion comprises at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, the ambulatory setting, affecting more than 15% of the US 50, 100, or more of the bacteria listed in Table X. In some population annually, resulting in over $5.8 billion in direct embodiments, the composition comprises at least one LAB health care expenditures (1). Typically classified by duration species. In some embodiments, the composition comprises of symptoms, sinusitis may be acute (less than 4 weeks in L. Sakei. In some embodiments, the composition consists duration), Sub-acute (4-12 weeks) or chronic (more than 12 essentially of L. Sakei, that is, other bacterial species, if weeks, with or without acute exacerbations). present at all, are not detectable, or not present in quantities 0004 Chronic Rhinosinusitis (CRS), represents a large Sufficient to colonize the sinonasal mucosa. portion of sinusitis cases, affecting more than 30 million 0008 Further provided are methods for improving the Americans (2), resulting in an annual economic health care sinus microbiota (e.g., increasing the microbial diversity, burden in excess of S2.4 billion. Culture-based studies have increasing the relative amount of beneficial bacteria Such as demonstrated chronic bacterial and/or fungal colonization of those listed in Table X, or reducing the level of pathogenic CRS patient sinus cavities implicating these species in the bacteria Such as Corynebacteria) in an individual comprising pathophysiology of CRS. Despite these findings, the micro administering at least one bacteria listed in Table X (e.g., a biology and immunology underlying CRS remains poorly Lactic Acid Bacterial (LAB) species, a Lactobacillus spe described, controversial, and to date no clear etiology has cies, or L. Sakei); and allowing the bacteria to colonize the been described (3). Known bacterial pathogens, such as sinonasal mucosa. Such methods can be used to treat an Staphylococcus and Streptococcus species isolated from individual diagnosed with sinusitis, or displaying sinusitis CRS sinuses and implicated in the disease (4-7), have also symptoms. In some embodiments, the administering is been detected in the nasopharynx of healthy individuals with nasal, e.g., using a spray, aerosol, Syringe, irrigation, or nasal no sinus symptomology (8). These results indicate that the drops. In some embodiments, the maxillary sinus is colo composition of the microbiota at discrete mucosal sites may nized. In some embodiments, the ethmoid sinus is colonized. define the abundance and pathogenic behavior of specific In some embodiments, the frontal sinus is colonized. In members of the assemblage. Such observations Suggest that Some embodiments, the sphenoid sinus is colonized. local microbiota composition plays a key role in protection 0009. In some embodiments, at least 2, 3, 4, 5, 6, 7, 8, 9. against pathogen overgrowth and virulence gene expression, 10, 20, 30, 40, 50, 100, or more of the bacterialisted in Table and that perturbations to local microbiota composition can X are administered. In some embodiments, at least one contribute to infectious and inflammatory disease etiology. Lactic Acid Bacterial (LAB) species is administered. In 0005. The sinuses are lined with respiratory epithelia that Some embodiments, Lactobacillus sakei is administered. In Support colonization by a diverse microbiome at upper Some embodiments, L. Sakei is the only bacteria adminis respiratory sites, e.g., oropharynx (8, 11). Little is known of tered in an effective amount (e.g., Sufficient to colonize the the composition of the resident microbiome of the paranasal sinonasal mucosa). sinuses and the contribution of these assemblages to sinus 0010. In some embodiments, the method further com mucosal health. Microbiological studies of this niche are prises detecting the microbial diversity of the sinonasal based on culture-based approaches, which under-estimate mucosa of the individual, before and/or after the adminis the diversity of species present. The present disclosure tering. In some embodiments, the method further comprises describes results from culture-independent approaches, and detecting the relative level of Corynebacteria (e.g., C. tuber thus provides a more accurate picture of sinus microbiome culostearicum and/or segmentosum) in a mucosal sample composition. The presently disclosed results show compara (e.g., sinonasal sample) from the individual and comparing tive analyses of healthy and diseased (sinusitis) samples, the relative level of Corynebacterium in the sample to a revealing both gross community characteristics and discrete control of Corynebacterium levels, before and/or after the species highly associated with health status. administering. US 2016/0310546 A1 Oct. 27, 2016

0011. Also provided are diagnostic methods, e.g., for bation to the sinus mucosal microbiota in the disease state. determining if an individual has or is at risk of developing FIG. 2D: Hierarchal cluster analysis demonstrates that the sinusitis. In some embodiments, the method of determining majority of healthy individuals cluster into a group distinct whether an individual has or is at risk of developing sinus from CRS patients, indicating that sinus microbiome com itis, comprises detecting the relative level of Corynebacteria position of healthy individuals is distinct form that of CRS in a mucosal sample from the individual; comparing the patients. A heat-plot of sinus symptom severity (based on relative level of Corynebacteria in the sample to a control of SNOT-20 scores) demonstrates that altered sinus microbi Corynebacteria levels; and determining that the individual ome composition is related to sinus health status. has or is at risk of developing sinusitis where the relative 0017 FIG. 3. Phylogenetic tree demonstrating the level of Corynebacteria in the sample is higher than the breadth of bacterial community members exhibiting signifi control, wherein the control is a non-sinusitis control, or in cant positive or negative correlations with sinus symptom the range of the control, wherein the control is a sinusitis severity (determined by SNOT-20 score). Corynebacterium positive control. In some embodiments, the mucosal sample tuberculostearicum and C. segmentosum exhibited Strong is from the maxillary sinus of the individual. In some and significant positive correlations with severity of sinus embodiments, the mucosal sample is from the ethmoid, symptomology, implicating them in disease etiology. Other frontal, or sphenoid sinus of the individual. In some embodi community members illustrated, e.g., in Table X, exhibited ments, the Corynebacteria is Corynebacterium tuberculoste significant negative correlations with sinus symptomology. aricum. The results indicate that these species maintain sinus 0012. In some embodiments, the method of determining mucosal health. whether an individual has or is at risk of developing sinus (0018 FIGS. 4A-4B. Linear regression of Q-PCR-derived itis, comprises detecting the relative level of at least one C. tuberculostearicum abundance with array reported flo bacteria from Table X (e.g., a LAB species, e.g., L. Sakei) in rescence intensity (FIG. 4A) and SNOT-20 score (FIG. 4B) a mucosal sample from the individual; comparing the rela indicate strong concordance between Q-PCR and array tive level of the at least one bacteria from Table X to a generated findings and confirm a strong positive relationship control of the level for the at least one bacteria; and between C. tuberculostearicum abundance and symptom determining that the individual has or is at risk of developing severity. sinusitis where the relative level of the at least one bacteria (0019 FIG. 5. Murine experimental design. from Table X in the sample is lower than the control, Amox=Amoxicillun; Clau=Clavulanate. wherein the control is a non-sinusitis control, or in the range 0020 FIGS. 6A-6B. FIG. 6A: PAS stained histological of the control, wherein the control is a sinusitis-positive sections of murine sinuses representative of animals in each control. In some embodiments, the mucosal sample is from treatment group treated with a combination of antibiotic and the maxillary sinus of the individual. In some embodiments, C. tuberculostearicum. Panel i-iv. Triplicate views of max the mucosal sample is from the ethmoid, frontal, or sphenoid illary sinuses from 2 mice per treatment group were used to sinus of the individual. In some embodiments, the at least determine physiology (representative images are shown). one bacteria is a LAB species, e.g., a Lactobacillus species. Mice treated with a combination of antibiotic and C. tuber 0013. In some embodiments, such methods further culostearicum show significantly increased goblet cell obtaining a mucosal sample from the sinonasal mucosa, e.g., hyperplasia (indicated by arrows) and mucin hypersecretion the maxillary sinus, of the individual before detecting. In compared to other treatment groups (panel iii). FIG. 6B: Some embodiments, a mucosal sample is taken from the Goblet cell enumeration illustrates that, compared to other ethmoid, frontal, or sphenoid sinus. In some embodiments, treatment groups, animals treated with a combination of the method is used to confirm a diagnosis of sinusitis using antibiotic and C. tuberculostearicum exhibit significant an alternative method, e.g., detection of elevated Muc5A increases in goblet cell number per um of epithelium. expression or mucosal inflammation, or a higher than nor (0021 FIG. 7. Q-PCR quantification of C. tuberculoste mal SNOT-20 score. arium and L. Sakei load in murine sinus tissue. Asterisk (*) 0014. In some embodiments, such methods further com denotes statistical significance (p<0.05). Total 16S rRNA prise administering a sinusitis therapeutic agent to the indi copy number per species was normalized to known number vidual based on a determination that the individual has or is of 16S rRNA copies per genome for each species. at risk of developing sinusitis. In some embodiments, the sinusitis therapeutic is a pharmaceutical composition com (0022 FIGS. 8A-8B. FIG. 8A: PAS stained histological prising at least one bacteria listed in Table X. In some sections of murine sinuses representative of animals in each embodiments, the pharmaceutical composition comprises at treatment group at 60x magnification (panel i-V). Animals treated with a combination of antibiotic and L. Sakei (panel least 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50 or more bacteria iv) exhibit epithelial physiology comparable to untreated or listed in Table X. In some embodiments, the pharmaceutical antibiotic treated controls (panels i and ii), while animals composition comprises a Lactic Acid Bacterial (LAB) spe treated with a combination of antibiotic and C. tuberculo cies, e.g., a Lactobacillus species such as L. Sakei. Stearicum exhibit goblet cell hyperplasia (indicated by BRIEF DESCRIPTION OF THE DRAWINGS arrows) and mucin hypersecretion. Panel V demonstrates that mice treated with antibiotics prior to co-instillation of C. 0015 FIG. 1. Muc5A mRNA expression (normalized to tuberculostearicum and L. Sakei exhibit epithelial physiol B-actin) is significantly increased in CRS patient samples ogy comparable to control animals, indicating that L. Sakei compared to healthy Subjects. is protective against the pathogenic effects of C. tuberculo 0016 FIGS. 2A-2D. Bacterial community Richness Stearicum. Triplicate views of maxillary sinuses from 2 mice (FIG. 2A), Evenness (FIG. 2B) and Shannon diversity (FIG. per treatment group were used to determine physiology 2C) indices are significantly lower in CRS patient sinuses (representative images are shown). FIG. 8B: Enumeration of compared with healthy individuals, indicating gross pertur goblet cells perum of epithelium confirms that hyperplasia US 2016/0310546 A1 Oct. 27, 2016

is associated with a high abundance of C. tuberculosteari illustrates why the etiology of CRS, and likely other chronic cum in the sinuses. Instillation of L. Sakei does not induce inflammatory diseases has been so difficult to define. hyperplasia, and co-instillation of L. Sakei with C. tubercu 0027. The role of the microbiome, LAB species in par lostearicum actually prevents the pathological response to ticular, in modulating the impact of C. tuberculostearicum C. tuberculostearicum. on sinus mucosal responses likely explains why it was not previously considered a pathogen. This phenomenon may DETAILED DESCRIPTION OF THE also explain why, despite detection of known pathogens in INVENTION the healthy subject sinus microbiota, these subjects exhibit no symptomology and, more broadly, provide an explana I. Introduction tion as to why seemingly similar patients, exhibiting com parable quantities of known pathogenic species, may exhibit 0023 Persistent mucosal inflammation and microbial dramatically different clinical outcomes. infection are characteristic of Chronic Rhinosinusitis (CRS). 0028. One advantage to the present comparative study The relationship between sinus microbiota composition and design is the ability to identify those species associated with CRS is unknown. The present disclosure describes compara healthy sinuses that provide mucosal protection. The murine tive microbiome profiling of a cohort of CRS patients and sinusitis model provided an opportunity to determine healthy Subjects, demonstrating that the sinus microbiota of whether Such species may afford protection against the CRS patients exhibit significantly reduced bacterial diver pathophysiology induced by a combination of depleted sity. Characteristic of this community collapse is the deple microbiota and C. tuberculostearicum. The results demon tion of multiple, phylogenetically distinct Lactic Acid Bac strate that L. Sakei represents a novel probiotic therapeutic teria and the concomitant increase in relative abundance of for the treatment of sinusitis sub-types, including CRS. Corynebacteria, in particular, C. tuberculostearicum. The Several members of the Lactobacillaceae, including L. Sakei, conditions of the human cohort were reproduced in a murine as well as other LABs and lactic acid producing members of model, and confirmed the pathogenic potential of C. tuber the Firmicutes were significantly depleted in CRS maxillary culostearicum, as well as the benefit of a diverse, properly sinus. The observation indicates that these species (e.g., composed mucosal microbiota to protect against C. tuber through bacteriocin or lactic acid production (30, 31)) can culostearicum. Moreover, the results show that Lactobacil out-compete pathogenic species, shape the sinus mucosal lus sakei, identified from comparative microbiome analyses microbiota, and protect this niche from pathogen over as protective, affords defense against C. tuberculostearicum growth. sinus infection, even in the context of a depleted sinus 0029. The present results also have significant implica bacterial community. The results demonstrate that sinus tions for the excessive use of antimicrobials, which contrib mucosal health depends on the local composition of resident ute to microbiota depletion, in the treatment of viral sinusitis microbiota, identifies a novel sinusitis associated pathogen, and other upper respiratory infections. The data Suggest that and a probiotic therapy for sinusitis. microbial Supplementation during periods of acute sinusitis 0024 Human health is dependent on the diverse micro with one or a combination of presently identified species bial assemblages that inhabit discrete host niches, particu would be more appropriate. larly mucosal-associated Surfaces. To date, culture-based approaches to characterize the etiological agent of CRS have II. Definitions provided a reductionist and somewhat discordant view of the microbiology associated with this disease. 0030 Unless defined otherwise, technical and scientific 0025. The culture-independent approaches described terms used herein have the same meaning as commonly herein allow the diversity of microbiomes in specific host understood by a person of ordinary skill in the art. See, e.g., sites to be better characterized (19-24), including composi Lackie, DICTIONARY OF CELL AND MOLECULARBIOLOGY, Elsevier tional and functional changes in disease States (25), and (4" ed. 2007); Sambrook et al., MOLECULAR CLONING, A microbes highly correlated with symptom severity (20) or LABORATORY MANUAL, Cold Springs Harbor Press (Cold immune responses (26). The present results show that the Springs Harbor, NY 1989). The term “a” or “an” is intended composition of the resident microbiota in a given niche can to mean “one or more.” The term “comprise' and variations strongly influence the behavior of specific species, particu thereof Such as “comprises and "comprising,” when pre larly a pathogen. ceding the recitation of a step or an element, are intended to 0026. A clear signal emerged from the present results mean that the addition of further steps or elements is demonstrating that the sinus microbiota of CRS patient optional and not excluded. Any methods, devices and mate cohort were characterized by both grossly depleted commu rials similar or equivalent to those described herein can be nities and a significant increase in relative abundance of C. used in the practice of this invention. The following defi tuberculostearicum. Though phylogenetically its closest nitions are provided to facilitate understanding of certain bacterial relatives include Mycobacteria and Nocardia, gen terms used frequently herein and are not meant to limit the era synonymous with pathogenesis, C. tuberculostearicum is Scope of the present disclosure. customarily considered an innocuous member of the healthy 0031. The term “colonize” in the context of the present skin microbiota, and an unlikely etiological agent of CRS. disclosure refers to microbial growth and expansion. For The presently described murine model of sinusitis confirmed example, a population of bacteria can be said to colonize an both the pathogenic potential of C. tuberculostearicum and environment if it survives in the environment through at that its impact on sinus epithelial responses was significantly least one round of cell division. enhanced in the absence of a replete sinus microbiota. 0032. The term “bacterial load’ refers to a measure of the Demonstration that a bacterial species that commonly inhab total number of bacteria in a given environment, e.g., an its human skin, represents an etiological agent of CRS airway mucosal Surface. The bacterial load is typically US 2016/0310546 A1 Oct. 27, 2016 expressed as colony forming units (CFU) per ml, per gram analyze databased on comparisons to control values. Con of sample or tissue, or per Surface area. trols are also valuable for determining the significance of 0033. The term “microbiome' refers to the community of data. For example, if values for a given parameter are widely microbes and environmental interactions in a defined envi variant in controls, variation in test samples will not be ronment. The microbiome also includes the genetic makeup considered as significant. of the microbes. Mucosal Surfaces are examples of micro 0040 “Subject,” “patient,” “individual” and like terms biomes in animals, e.g., the gut, airways, sinus, Vagina, oral are used interchangeably and refer to, except where indi cavity, etc. Skin, and particular skin areas, comprise addi cated, humans and non-human animals. The term does not tional microbiomes. The term is derived from the term necessarily indicate that the Subject has been diagnosed with “biome' which can be used synonymously with ecosystem a particular disease, but typically refers to an individual to refer to a larger scale environment and its inhabitants. under medical Supervision. A patient can be an individual 0034. The term “improving the sinus microbiota refers that is seeking treatment, monitoring, adjustment or modi to increasing microbial diversity, increasing the relative fication of an existing therapeutic regimen, etc. amount of beneficial bacteria (e.g., bacteria listed in Table 0041. The term “in the range of the control refers to a X), or reducing the relative amount of bacteria with high value that falls within the range of control values for a given levels in sinusitis patients (e.g., Corynebacteria). control condition. Where one value is given as a control, “in 0035 Microbial diversity refers to the range of different the range of the control” refers to a value that is not species or strains present in a sample. The sample can be statistically different from the control as determined by one highly diverse, with a comparatively wide range of taxa, or of skill in the art. For example, within the range of the lacking in diversity, e.g., with comparatively few taxa rep control can be +5%, 10%, 20%, or within 0.5-fold, 1-fold, or resented compared to a normal control. Microbial diversity 2-fold difference from a control value at a given condition. can be expressed in general comparative terms (e.g., more or One of skill will understand that corrective calculations can less diverse, compared to a normal, healthy control). Micro be made, e.g., to account for age, severity of condition, bial diversity can also be expressed in absolute numbers or antibiotic use, etc. ranges of numbers, e.g., more than 1000 species or strains, 0042. As used herein, the term “pharmaceutically accept or 500-100, 50-100 or, 25-50 species or strains, etc. in a able' is used synonymously with physiologically acceptable given environment. and pharmacologically acceptable with respect to, e.g., a 0036. The amount of a particular strain or species of pharmaceutical composition. A pharmaceutical composition bacteria can be expressed in absolute numbers or ranges of will generally comprise agents for buffering and preserva numbers, e.g., 10° or 10°-107 Lactobacillus in a sample, or tion in storage, and can include buffers and carriers for in a given volume. The amount can also be expressed in appropriate delivery, depending on the route of administra terms of colony forming units, or absorbance, depending on tion. the assay used for bacterial detection, as will be appreciated 0043. The terms “dose' and “dosage' are used inter by one of skill. The amount can also be expressed in changeably herein. A dose refers to the amount of active comparative terms, e.g., compared to a control. The relative ingredient given to an individual at each administration. For level of a given strain or species refers to the amount relative the present invention, the dose will generally refer to the to other strains or species within the sample. For example, amount of probiotic, antibiotic or anti-inflammatory agent. a high relative level of a Corynebacterial strain indicates that Dosage can also be expressed in terms of bacterial concen the amount of Corynebacterial strain, as a percentage of total tration. The dose will vary depending on a number of microbes, is higher than that of a normal control. This can factors, including frequency of administration; size and be due to depletion of other species in the microbiome, or tolerance of the individual; severity of the condition; risk of because of high levels of Corynebacteria. side effects; and the route of administration. One of skill will 0037 “Lactic acid bacteria” (LAB) refers to species that recognize that the dose can be modified depending on the fall into the order of Gram-positive Lactobacillalese. LAB above factors or based on therapeutic progress. The term genera include: Lactobacillus, Leuconostoc, Pediococcus, “dosage form” refers to the particular format of the phar Lactococcus, and Streptococcus, as well as Aerococcus, maceutical, and depends on the route of administration. For Carnobacterium, Enterococcus, and Tetragenococcus. example, a dosage form can be in a liquid form for nebu 0038 A “control sample or value refers to a sample that lization, e.g., for inhalants, in a tablet or liquid, e.g., for oral serves as a reference, usually a known reference, for com delivery, or a saline solution, e.g., for injection. parison to a test sample. For example, a test sample can be 0044 As used herein, the terms “treat' and “prevent are taken from a patient Suspected of having sinusitis and not intended to be absolute terms. Treatment can refer to any compared to samples from a known sinusitis patient (posi delay in onset, reduction in the frequency or severity of tive or disease control) or a known normal (negative, non symptoms, amelioration of symptoms, improvement in sinusitis, non-disease, or healthy control) individual. A patient comfort and/or respiratory function, etc. The effect of control can also represent an average range or value gath treatment can be compared to an individual or pool of ered from a population of similar individuals, e.g., sinusitis individuals not receiving a given treatment, or to the same patients or healthy individuals with a similar medical back patient prior to, or after cessation of treatment. ground, same age, weight, etc. A control value can also be 0045. The term “prevent” refers to a decrease in the obtained from the same individual, e.g., from an earlier occurrence of sinusitis symptoms in a patient. As indicated obtained sample, prior to disease, or prior to treatment. One above, the prevention may be complete (no detectable of skill will recognize that controls can be designed for symptoms) or partial. Such that fewer symptoms are assessment of any number of parameters. observed than would likely occur absent treatment. 0039. One of skill in the art will understand which 0046. The terms “effective amount” and “therapeutically controls are valuable in a given situation and be able to effective amount,” refer to that amount of a therapeutic agent US 2016/0310546 A1 Oct. 27, 2016

sufficient to ameliorate the target disorder. For example, for analogs may have modified R groups (e.g., norleucine) or a given disease parameter, a therapeutically effective amount modified peptide backbones, but retain the same basic will show an increase or decrease of at least 5%, 10%, 15%, chemical structure as a naturally occurring amino acid. 20%, 25%, 40%, 50%, 60%, 75%, 80%, 90%, or at least Amino acid mimetics refers to chemical compounds that 100%. Therapeutic efficacy can also be expressed as “-fold' have a structure that is different from the general chemical increase or decrease. For example, a therapeutically effec structure of an amino acid, but that functions similarly to a tive amount can have at least a 1.2-fold, 1.5-fold, 2-fold, naturally occurring amino acid. 5-fold, or more effect over a control. 0051 Amino acids may be referred to herein by either 0047. The term “diagnosis” refers to a relative probability their commonly known three letter symbols or by the that a disease is present in the subject. Similarly, the term one-letter symbols recommended by the IUPAC-IUB Bio “prognosis' refers to a relative probability that a certain chemical Nomenclature Commission. Nucleotides, likewise, future outcome may occur in the Subject. The terms are not may be referred to by their commonly accepted single-letter intended to be absolute, as will be appreciated by one of skill codes. in the field of medical diagnostics. 0.052 “Conservatively modified variants' applies to both 0048 “Nucleic acid” or “oligonucleotide' or “polynucle amino acid and nucleic acid sequences. With respect to otide' or grammatical equivalents used herein means at least particular nucleic acid sequences, conservatively modified two nucleotides covalently linked together. Oligonucle variants refers to those nucleic acids which encode identical otides are typically from about 5, 6, 7, 8, 9, 10, 12, 15, 25, or essentially identical amino acid sequences, or where the 30, 40, 50 or more nucleotides in length, up to about 100 nucleic acid does not encode an amino acid sequence, to nucleotides in length. Nucleic acids and polynucleotides are essentially identical or associated, e.g., naturally contiguous, a polymers of any length, including longer lengths, e.g., 200, sequences. Because of the degeneracy of the genetic code, a 300, 500, 1000, 2000, 3000, 5000, 7000, 10,000, etc. A large number of functionally identical nucleic acids encode nucleic acid of the present invention will generally contain most proteins. For instance, the codons GCA, GCC, GCG phosphodiester bonds, although in some cases, nucleic acid and GCU all encode the amino acid alanine. Thus, at every analogs are included that may have alternate backbones, position where an alanine is specified by a codon, the codon comprising, e.g., phosphoramidate, phosphorothioate, phos can be altered to another of the corresponding codons phorodithioate, or O-methylphophoroamidite linkages (see described without altering the encoded polypeptide. Such Eckstein, Oligonucleotides and Analogues: A Practical nucleic acid variations are “silent variations,” which are one Approach, Oxford University Press); and peptide nucleic species of conservatively modified variations. Every nucleic acid backbones and linkages. Other analog nucleic acids acid sequence herein which encodes a polypeptide also include those with positive backbones; non-ionic backbones, describes silent variations of the nucleic acid. One of skill and non-ribose backbones, including those described in U.S. will recognize that in certain contexts each codon in a Pat. Nos. 5,235,033 and 5,034,506, and Chapters 6 and 7. nucleic acid (except AUG, which is ordinarily the only ASC Symposium Series 580, Carbohydrate Modifications in codon for methionine, and TGG, which is ordinarily the only Antisense Research, Sanghui & Cook, eds. Nucleic acids codon for tryptophan) can be modified to yield a function containing one or more carbocyclic Sugars are also included ally identical molecule. Accordingly, often silent variations within one definition of nucleic acids. Modifications of the of a nucleic acid which encodes a polypeptide is implicit in ribose-phosphate backbone may be done for a variety of a described sequence with respect to the expression product, reasons, e.g., to increase the stability and half-life of Such but not with respect to actual probe sequences. molecules in physiological environments or as probes on a 0053 As to amino acid sequences, one of skill will biochip. Mixtures of naturally occurring nucleic acids and recognize that individual Substitutions, deletions or addi analogs can be made; alternatively, mixtures of different tions to a nucleic acid, peptide, polypeptide, or protein nucleic acid analogs, and mixtures of naturally occurring sequence which alters, adds or deletes a single amino acid or nucleic acids and analogs may be made. a small percentage of amino acids in the encoded sequence 0049. The terms “polypeptide,” “peptide' and “protein' is a “conservatively modified variant' where the alteration are used interchangeably herein to refer to a polymer of results in the Substitution of an amino acid with a chemically amino acid residues. The terms apply to amino acid poly similar amino acid. Conservative substitution tables provid mers in which one or more amino acid residue is an artificial ing functionally similar amino acids are well known in the chemical mimetic of a corresponding naturally occurring art. Such conservatively modified variants are in addition to amino acid, as well as to naturally occurring amino acid and do not exclude polymorphic variants, interspecies polymers, those containing modified residues, and non homologs, and alleles of the invention typically conservative naturally occurring amino acid polymer. substitutions for one another: 1) Alanine (A), Glycine (G); 0050. The term "amino acid refers to naturally occurring 2) Aspartic acid (D), Glutamic acid (E); 3) Asparagine (N), and synthetic amino acids, as well as amino acid analogs and Glutamine (Q); 4) Arginine (R), Lysine (K); 5) Isoleucine amino acid mimetics that function similarly to the naturally (I), Leucine (L), Methionine (M), Valine (V); 6) Phenylala occurring amino acids. Naturally occurring amino acids are nine (F), Tyrosine (Y), Tryptophan (W); 7) Serine (S), those encoded by the genetic code, as well as those amino Threonine (T); and 8) Cysteine (C), Methionine (M) (see, acids that are later modified, e.g., hydroxyproline, Y-car e.g., Creighton, Proteins (1984)). boxyglutamate, and O-phosphoserine. Amino acid analogs 0054) The terms “label,” “tag” “detectable moiety,” etc. refers to compounds that have the same basic chemical refer to compositions or components that are detectable by structure as a naturally occurring amino acid, e.g., an a spectroscopic, photochemical, biochemical, immunochemi carbon that is bound to a hydrogen, a carboxyl group, an cal, chemical, or other physical means. For example, useful amino group, and an R group, e.g., homoserine, norleucine, labels include ’P. fluorescent dyes, electron-dense reagents, methionine sulfoxide, methionine methyl sulfonium. Such enzymes (e.g., as commonly used in an ELISA), biotin, US 2016/0310546 A1 Oct. 27, 2016 digoxigenin, or haptens and proteins or other entities which 0059. The term “antibody” refers to a polypeptide struc can be made detectable, e.g., by incorporating a radiolabel ture, e.g., an immunoglobulin, conjugate, or fragment into a peptide or antibody specifically reactive with a target thereof that retains antigen binding activity, e.g., for a peptide. Any method known in the art for conjugating an bacterial antigen. The term includes but is not limited to antibody to the label may be employed, e.g., using methods polyclonal or monoclonal antibodies of the isotype classes described in Hermanson, Bioconjugate Techniques 1996, IgA, Ig|D, IgE, IgG, and IgM, derived from human or other Academic Press, Inc., San Diego. mammalian cells, including natural or genetically modified 0055. A “labeled nucleic acid probe or oligonucleotide' forms such as humanized, human, single-chain, chimeric, is one that is bound, either covalently, through a linker or a synthetic, recombinant, hybrid, mutated, grafted, and in chemical bond, or noncovalently, through ionic, van der vitro generated antibodies. The term encompases conju Waals, electrostatic, or hydrogen bonds to a label such that gates, including but not limited to fusion proteins containing the presence of the probe may be detected by detecting the an immunoglobulin moiety (e.g., chimeric or bispecific presence of the label bound to the probe. Alternatively, antibodies or schv's), and fragments, such as Fab, F(ab')2. method using high affinity interactions may achieve the Fv, scFv, Fd, dAb and other compositions. same results where one of a pair of binding partners binds to 0060 An exemplary immunoglobulin (antibody) struc the other, e.g., biotin, Streptavidin. tural unit comprises a tetramer. Each tetramer is composed of two identical pairs of polypeptide chains, each pair having 0056. The phrase “selectively (or specifically) hybridizes one “light” (about 25 kD) and one “heavy chain (about to refers to the binding, duplexing, or hybridizing of a 50-70 kD). The N-terminus of each chain defines a variable molecule only to a particular nucleotide sequence with a region of about 100 to 110 or more amino acids primarily higher affinity, e.g., under more stringent conditions, than to responsible for antigen recognition. Typically, the “variable other nucleotide sequences (e.g., total cellular or library region' contains the antigen-binding region of the antibody DNA or RNA). One of skill in the art will appreciate that (or its functional equivalent) and is most critical in speci specific hybridization between nucleotides usually relies on ficity and affinity of binding. See Paul, Fundamental Immu Watson-Crick pair bonding between complementary nucleo nology (2003). The terms variable light chain (V) and tide sequences. variable heavy chain (V) refer to these light and heavy 0057 The term “probe' or “primer', as used herein, is chains respectively. defined to be one or more nucleic acid fragments whose 0061 Antibodies can exist as intact immunoglobulins or specific hybridization to a sample can be detected. A probe as any of a number of well-characterized fragments pro or primer can be of any length depending on the particular duced by digestion with various peptidases. Pepsin digests technique it will be used for. For example, PCR primers are an antibody below the disulfide linkages in the hinge region generally between 10 and 40 nucleotides in length, while to produce F(ab)', a dimer of Fab which itself is a light nucleic acid probes for, e.g., a Southern blot, can be more chain joined to V-C1 by a disulfide bond. The F(ab)' may than a hundred nucleotides in length. The probe may be be reduced under mild conditions to break the disulfide unlabeled or labeled as described below so that its binding linkage in the hinge region, thereby converting the F(ab)' to the target or sample can be detected. The probe can be dimer into an Fab' monomer. The Fab' monomer is essen produced from a source of nucleic acids from one or more tially Fab with part of the hinge region (see Fundamental particular (preselected) portions of a chromosome, e.g., one Immunology (Paul ed., 3d ed. 1993). While various antibody or more clones, an isolated whole chromosome or chromo fragments are defined in terms of the digestion of an intact Some fragment, or a collection of polymerase chain reaction antibody, one of skill will appreciate that such fragments (PCR) amplification products. The length and complexity of may be synthesized de novo either chemically or by using the nucleic acid fixed onto the target element is not critical recombinant DNA methodology. Thus, the term antibody, as to the invention. One of skill can adjust these factors to used herein, also includes antibody fragments either pro provide optimum hybridization and signal production for a duced by the modification of whole antibodies, or those given hybridization procedure, and to provide the required synthesized de novo using recombinant DNA methodologies resolution among different genes or genomic locations. (e.g., single chain FV) or those identified using phage display 0058. The probe may also be isolated nucleic acids libraries (see, e.g., McCafferty et al., Nature 348:552-554 immobilized on a Solid Surface (e.g., nitrocellulose, glass, (1990)). quartz, fused silica slides), as in an array. In some embodi 0062. As used herein, “V-region” refers to an antibody ments, the probe may be a member of an array of nucleic variable region domain comprising the segments of Frame acids as described, for instance, in WO 96/17958. Tech work 1, CDR1, Framework 2, CDR2, and Framework 3, niques capable of producing high density arrays can also be including CDR3 and Framework 4, which segments are used for this purpose (see, e.g., Fodor (1991) Science added to the V-segment as a consequence of rearrangement 767-773: Johnston (1998) Curr. Biol. 8: R171-R174; of the heavy chain and light chain V-region genes during Schummer (1997) Biotechniques 23: 1087-1092: Kern B-cell differentiation. (1997) Biotechniques 23: 120-124; U.S. Pat. No. 5,143,854). 0063 As used herein, "complementarity-determining One of skill will recognize that the precise sequence of the region (CDR) refers to the three hypervariable regions in particular probes described herein can be modified to a each chain that interrupt the four “framework” regions certain degree to produce probes that are “substantially established by the light and heavy chain variable regions. identical to the disclosed probes, but retain the ability to The CDRs are primarily responsible for binding to an specifically bind to (i.e., hybridize specifically to) the same epitope of an antigen. The CDRS of each chain are typically targets or samples as the probe from which they were referred to as CDR1, CDR2, and CDR3, numbered sequen derived. Such modifications are specifically covered by tially starting from the N-terminus, and are also typically reference to the individual probes described herein. identified by the chain in which the particular CDR is US 2016/0310546 A1 Oct. 27, 2016

located. Thus, a V. CDR3 is located in the variable domain of various sample gathering techniques is described, e.g., in of the heavy chain of the antibody in which it is found, Roediger et al. (2010) Am J Rhinol Allergy 24:263. whereas a V, CDR1 is the CDR1 from the variable domain 0069. A diagnostic agent specific for the microbes of the light chain of the antibody in which it is found. described herein can include any label as known in the art, 0064. The terms “antigen,” “immunogen,” “antibody tar for example, in the following references: Armstrong et al., get,” “target analyte.” and like terms are used herein to refer Diagnostic Imaging, 5' Ed., Blackwell Publishing (2004); to a molecule, compound, or complex that is recognized by Torchilin, V. P. Ed., Targeted Delivery of Imaging Agents, an antibody, i.e., can be specifically bound by the antibody. CRC Press (1995); Vallabhajosula, S., Molecular Imaging: The term can refer to any molecule that can be specifically Radiopharmaceuticals for PET and SPECT. Springer recognized by an antibody, e.g., a polypeptide, polynucle (2009). A diagnostic agent can be detected by a variety of otide, carbohydrate, lipid, chemical moiety, or combinations ways, including as an agent providing and/or enhancing a thereof (e.g., phosphorylated or glycosylated polypeptides, detectable signal. Detectable signals include, but are not etc.). One of skill will understand that the term does not limited to, fluorescent, luminescent, gamma-emitting, radio indicate that the molecule is immunogenic in every context, active, optical, absorptive, etc. The terms “detectable agent,” but simply indicates that it can be targeted by an antibody. “detectable moiety.” “label,” “imaging agent, and like Examples of target antigens in the context of the present terms are used synonymously herein. disclosure include bacterial antigens (e.g., from C. tubercu 0070 A radioisotope can be incorporated into the diag lostearicum, L. Sakei, or an LAB species) and disease nostic agents described herein and can include radionuclides associated antigens, e.g., Muc5A. that emit gamma rays, positrons, beta and alpha particles, 0065 Antibodies bind to an “epitope' on an antigen. The and X-rays. Suitable radionuclides include but are not lim epitope is the localized site on the antigen that is recognized ited to 225Ac, 7°As, 21 At 1B, 12 Ba, 212Bi, 75 Br, 77Br, 14C, and bound by the antibody. Epitopes can include a few 10°Cd, 62Cu, Cu, 7Cu, 18F 7Ga, Ga, H, 16Ho, 123, amino acids or portions of a few amino acids, e.g., 5 or 6. 124, 125I, 130I, 131I, 111 In 177Lu, 13N, 15O, 32P 33P. 212Pb, or more, e.g., 20 or more amino acids, or portions of those 105Pd, 18.Re, 18.Re, 7Sc, 'Sim, 89Sr. 99mTc, 88Y and 90Y. amino acids. In some cases, the epitope includes non-protein 0071. In some embodiments, the diagnostic agent can be components, e.g., from a carbohydrate, nucleic acid, or lipid. associated with a secondary binding ligand or to an enzyme In some cases, the epitope is a three-dimensional moiety. (an enzyme tag) that will generate a colored product upon Thus, for example, where the target is a protein, the epitope contact with a chromogenic Substrate. Examples of Suitable can be comprised of consecutive amino acids, or amino enzymes include urease, alkaline phosphatase, (horseradish) acids from different parts of the protein that are brought into hydrogen peroxidase and glucose oxidase. Secondary bind proximity by protein folding (e.g., a discontinuous epitope). ing ligands include, e.g., biotin and avidin or streptavidin The same is true for other types of target molecules that form compounds as known in the art. three-dimensional structures. 0072. In some embodiments, the diagnostic agents can include optical agents such as fluorescent agents, phospho III. Diagnostic Methods rescent agents, chemiluminescent agents, and the like. 0066. The present results reveal that sinusitis patients Numerous agents (e.g., dyes, probes, labels, or indicators) have greatly reduced microbial diversity in the sinonasal are known in the art and can be used in the present invention. environment (e.g., maxillary sinus). In addition, Corynebac (See, e.g., Invitrogen, The Handbook A Guide to Fluores terial species (e.g., C. tuberculostearicum and segmentosum) cent Probes and Labeling Technologies, Tenth Edition are present at a higher level than in normal healthy sinus (2005)). Fluorescent agents can include a variety of organic mucosa, and correlate strongly with symptom severity. and/or inorganic Small molecules or a variety of fluorescent Sinusitis can also be diagnosed, e.g., by detecting mucin proteins and derivatives thereof. For example, fluorescent levels, mucosal inflammation, or with self-reporting of sinus agents can include but are not limited to cyanines, phthalo inflammation symptoms. Diagnostic methods can be used cyanines, porphyrins, indocyanines, rhodamines, phenox alone or in any combination. azines, phenylxanthenes, phenothiazines, phenoselenazines, 0067 Patients can thus be selected for therapy based on fluoresceins, benzoporphyrins, squaraines, dipyrrolo pyrimi determination of microbial diversity, reduced levels of ben dones, tetracenes, quinolines, pyrazines, corrins, croconi eficial probiotic species (e.g., those listed in Table X, Lactic ums, acridones, phenanthridines, rhodamines, acridines, Acid Bacteria, or L. sakei in particular), or elevated levels of anthraquinones, chalcogenopyrylium analogues, chlorins, Corynebacteria. Anatomical characterization, such as char naphthalocyanines, methine dyes, indolenium dyes, azo acterization of the sinus mucosal lining, can be accom compounds, aZulenes, azaaZulenes, triphenyl methane dyes, plished using standard imaging techniques. Bacteria can be indoles, benzoindoles, indocarbocyanines, benzoindocarbo detected using nucleic acid techniques as described herein cyanines, and BODIPYTM derivatives. (e.g., arrays, hybridization, or PCR), using sequences complementary to species- or order-specific nucleic acid IV. Methods and Compositions for Administration sequences. Antibodies specific for particular bacterial spe 0073 Provided herein are methods of improving sinus cies can also be utilized. microbiota (e.g., increasing microbial diversity or reducing 0068 Methods of obtaining a mucosal sample from an the relative population of pathogenic species) in an indi individual are known in the art. Such methods include vidual comprising administering a pharmaceutical compo Swabbing or brushing the sinonasal mucosa, e.g., using sition comprising a beneficial microbe, e.g., from Table X, anesthetic or endoscopic methods if necessary. In some to the individual. Typically, the composition is administered cases, a mucosal biopsy is taken. Samples can also be nasally, trans-nasally, or to the sinuses, e.g., using an aero obtained using a nasal lavage or spray in Sufficient volume Sol, spray, irrigation, or nasal drops. Such methods can be to obtain sample from the appropriate location. Comparison used to treat an individual diagnosed with sinusitis. US 2016/0310546 A1 Oct. 27, 2016

0.074. A pharmaceutical composition comprising a ben goblet cell hyperplasia, or increased microbial diversity). In eficial microbe described herein can be administered, alone Some cases, however, the initial dose is relatively high to or in combination with other Suitable components, using an establish a colonizing population in a patient experiencing aerosol formulations (“nebulized') to be administered via acute symptoms. One of skill will appreciate that a number inhalation. Aerosol formulations can be placed into pressur of variables must be considered when determining a thera ized acceptable propellants, such as dichlorodifluorometh peutically effective dose. The dose administered to a patient ane, propane, nitrogen, etc. should be sufficient to effect a beneficial therapeutic 0075 Compositions for administration typically com response in the patient over time. The size of the dose also prises at least one probiotic microbe, e.g., as identified in will be determined by the existence, nature, and extent of Table X, in a pharmaceutically acceptable carrier, e.g., an any adverse side-effects that accompany the administration aqueous carrier. A variety of aqueous carriers can be used, of a particular composition (or combination therapy) in a e.g., buffered saline. These solutions are generally free of particular patient. undesirable matter, e.g., contaminating species. The com positions can contain pharmaceutically acceptable auxiliary V. Kits Substances as required to approximate physiological condi I0081 Further provided are kits for detecting at least one tions such as pH adjusting and buffering agents, for example, sinusitis associated bacterial taxa (i.e., bacteria with high Sodium acetate, sodium chloride, potassium chloride, cal levels correlating with sinusitis, or bacteria are associated cium chloride, Sodium lactate, etc. The concentration of with the absence of sinusitis, such as the bacteria in Table active agent in these formulations can vary widely, and will X). The kit can optionally include written instructions, be selected primarily based on fluid volumes, viscosities, reference to an internet site, or electronic instructions (e.g., body weight and the like in accordance with the particular on a CD-ROM or DVD). In some embodiments, kits of the mode of administration selected and the patient’s needs. invention will include a case or container for holding the 0076. The formulation may also provide additional active reagents in the kit, which can be included separately or in compounds, including, e.g., antibiotic or anti-inflammatory combination. In some embodiments, the kit includes agents. Combination therapies contemplate coadministra brushes, Swabs, or other sample gathering devices for tion, using separate formulations or a single pharmaceutical obtaining a sinonasal sample from an individual, e.g., from formulation, and consecutive administration in either order. the maxillary, ethmoid, frontal, and/or sphenoid sinus. The 0077. The active ingredients can be prepared as sus kit can further comprise sample containers for holding and tained-release preparations (e.g., semi-permeable matrices processing samples, and for detection of sinonasal microbes of Solid hydrophobic polymers (e.g., polyesters, hydrogels in the sample. (for example, poly (2-hydroxyethyl-methacrylate), or poly I0082 In some embodiments, the kit includes an array, the (vinylalcohol)), polylactides. The probiotic formulations array comprising probes or other agents capable of specific described herein can also be entrapped in microcapsules detection of at least one sinusitis associated bacterial taxa. prepared, for example, by coacervation techniques or by For example, the array can comprise probes that specifically interfacial polymerization, for example, hydroxymethylcel hybridize to nucleotide sequences from the at least one lulose or gelatin microcapsules and poly-(methylmethacy sinusitis associated bacteria, e.g., polynucleotide or oligo late) microcapsules, respectively, in colloidal drug delivery nucleotide probes. In some embodiments, the array com systems (for example, liposomes, albumin microspheres, prises components (e.g., antibody fragments) that specifi microemulsions, nano-particles and nanocapsules) or in cally detect non-nucleic acid markers from the at least one macroemulsions. sinusitis associated bacteria. In some embodiments, the 0078 Methods for preparing therapeutic compositions array can specifically detect at least 4, 5, 10, 12, 15, 20, 50. will be known to those skilled in the art and are described in 80, 100, or all of the sinusitis associated bacteria, e.g., more detail in Such publications as Remington's Pharma Corynebacteria or those listed in Table X. In some embodi ceutical Science, 15th ed., Mack Publishing Company, Eas ments, the kit further includes reagents (buffers, secondary ton, Pa. (1980). detection agents, etc.) required for running the detection 007.9 The compositions can be administered for thera reaction. peutic or prophylactic treatments. In therapeutic applica 0083. In some embodiments, the kit will include PCR tions, compositions are administered to a patient Suffering reagents and primers for detecting at least one of the bacteria from a disease (e.g., sinusitis) in an effective dose. Amounts disclosed in Table X, or at least one Corynebacterial species. effective for this use will depend upon the route of admin The kit can thus include a polymerase, nucleotide mono istration, the severity of the condition, and the general state mers, buffer stocks, and optionally an interchalating fluo of the patient’s health. Single or multiple administrations of rescent dye. In some embodiments, the kit can include PCR the compositions may be administered depending on the primers for at least 4, 5, 10, 12, 15, 20, 50, 60, or all of the dosage and frequency as required and tolerated by the sinusitis associated bacteria, e.g., Corynebacteria or those patient. The presently described compositions can be admin listed in Table X. istered to humans and other animals. Thus the methods are 0084. In some embodiments, the kit can include PCR applicable to both human therapy and Veterinary applica primers for detecting a polynucleotide from at least one tions. sinusitis associated bacteria, e.g., Corynebacteria (e.g., 0080. To determine a therapeutically effective dose, e.g. tuberculostearicum or segmentosum), or at least one bacteria a colonizing dose of a probiotic composition, a relatively listed in Table X. The primers can be designed to amplify low dose of the composition can be initially administered to more than one species or Strain from a bacterial taxa, e.g., the individual, and the dose can be incrementally increased more than one Corynebacteria or Lactobacillus strain. until the condition of the individual, e.g., the sinus environ I0085. In some embodiments, the kit will include a posi ment, begins to improve (e.g., reduced mucin secretion or tive control for the at least one sinusitis associated bacteria US 2016/0310546 A1 Oct. 27, 2016

(e.g., an isolated Sample of the bacteria). In some embodi change tree using FastTree (43) which was annotated using ments, the kit will include a negative control for the at least the Interactive Tree of Life website accessible at itol.embl.de one sinusitis associated bacteria (e.g., a blank, or an unre (44). lated bacterial sample). 0092 Murine Sinus Studies. I0086. It is understood that the examples and embodi (0093. Female 4-5 week old C57BL6/J mice weighing 16 ments described herein are for illustrative purposes only and to 18 g were purchased form Jackson Laboratories (Bar that various modifications or changes in light thereof will be Harbor, Me.) and housed in micro-isolator cages. Animals Suggested to persons skilled in the art and are to be included were permitted to acclimatize for two weeks with food and within the spirit and purview of this application and scope of water ad libitum prior to study. Mice were administered the appended claims. All publications, patents, patent appli augmentin (amoxicillin/clavulanate) at 100 mg/kg dosage cations, Accession or ID numbers, and websites cited herein once a day for five days prior to bacterial instillation. Prior are hereby incorporated by reference in their entirety for all to intra-nasal C. tuberculostearicum, L. Sakei, or combina purposes. tion of both C. tuberculostericum and L. Sakei, inoculations mice were anesthetized by intraperitoneal injection with VI. Examples avertin (250 mg/kg). Inoculation was performed once a day for three days by applying 25ul of either C. tuberculsteari A. Materials and Methods cum suspension (1.0x10' cfu/ml) or L. sakei (1.0x10' cfu/ml) suspension, or combination of both microbes in 0087 Patient Sample Collection. equal ratios (total 2.0x10' cfu/ml), in PBS onto the external 0088 Patient and disease stratification was performed nares and inhalation by the animals (see e.g., 45, 46). Mice based on recent clinical history, nasal endoscopy, CT sinus were monitored for breathing during the entire inoculation review, and a validated quality-of-life instrument, the dis process and post inoculation until fully recovered from ease-specific Sinonasal Outcome Test survey SNOT-20; anesthesia. Twenty-four hours after the final bacterial inocu (38). Sinus brushings were obtained during functional lation, mice were euthanized by CO asphyxiation followed endoscopic sinus surgery (FESS) of CRS patients, or surgery by induced pneumothorax. The heads were decapitated and for non-CRS complaints, e.g., obstructive sleep apnea or sinuses dissected for histological (n=2 animals per treatment post-traumatic malocclusion, in healthy Subjects. Endo group) and molecular analysis (n=3 animals per treatment scopically-guided brush samples of mucosal Surfaces of the group). Dissected sinuses used for molecular analyses were lateral, central, and medial portions of the maxillary sinus placed in RNAlater (Ambion, TX) and stored at 4°C. until were obtained and pooled together in 1 ml of RNALater for processed the following day in the same manner as analysis. Samples were placed at 4°C. for 24 hours prior to described above for human samples. storage at -80° C. until processed. (0094 Sinus Histology. 0.095 Sinuses used for histological analysis were fixed I0089 DNA Extraction and PhyloChip Analysis. overnight in 4% paraformaldehye, followed by overnight 0090. Mucosal brushings were transferred to Lysis decalcification in Decalcifying solution A (Fisher, CA). Matrix B (MP Biomedicals, OH) tubes containing 600 ul Samples were then dehydrated as follows: rinsed with PBS RLT buffer (Qiagen, CA). Samples were subjected to 30 (1 hour), soaked in 30% ethanol (1 hour), 50% ethanol (1 seconds of bead beating at 5.5 m/second followed by cen hour), then transferred to 70% ethanol and stored at 4°C. trifugation for one minute at 2000 rpm. Supernatant was until Subsequent preparation. Sinus Samples were sectioned transferred to the AllPrepDNA spin column and nucleic acid to 5um thickness and mounted onto glass microscope slides. purification (DNA and RNA) was carried out according to H&E and PAS staining were performed as previously the manufacturer's instructions (Qiagen, CA). Nucleic acid described (47). PAS-stained sections prepared from four concentrations were determined using a NanoDrop specto groups of mice (CTAB-, CT-AB+, CT+AB- and CT+AB-) photometer (Thermo Scientific, DE). PhyloChip analysis were imaged at 20x and 60x magnifications, and PAS was performed as previously described (40) using 250 ng of positive cells were counted for 3 different sections per mice purified pooled 16S rRNA amplicon per sample generated (2 mice in each group). The number of goblet cells was using 27F and 1492R universal primers (41). Data sets were expressed as the total number of PAS-positive cells per um conservatively filtered as previously described (40). Probe length of epithelium. Students t-test was used to calculate set fluorescence intensity was normalized and log trans p-values, p<0.05 was considered significant. formed prior to analysis using packages in the R statistical 0096 Q-PCR. environment. Hierarchical cluster analysis (HCA) was per 0097 Bacterial burden was determined using extracted formed on a Bray-Curtis dissimilarity matrix generated from DNA (10 ng per sample, triplicate reactions) and universal PhyloChip fluorescence intensity data using the vegan pack 16S rRNA Q-PCR primers 338F, 5'-ACTC age as we have previously described (22), followed by CTACGGGAGG CAGCAG-3 (41) and 518R 5'-ATTAC average linkage clustering. CGCGGCTGCTGG-3'(48). Quantitec SYBR Green (Qia 0091. A two-tailed Welch's T-test was used to identify gen, CA) was used according to manufacturers instructions. taxa that were significantly altered in relative abundance Reaction mixtures (25 ul total) contained 12.5 ul of 2x across healthy Subject and patient groups and adjusted for QuantiTec SYBR green (Qiagen, CA), 2.5 ul each of 3 uM false discovery (42) using the q-value package as previously forward and reverse primer, and 6.5 ul H.O. Reactions were described at the website accessible at cran.r-project.org/web/ amplified using the MX3000P Real-Time PCR System packages/qvalue/qvalue.pdf. Results with a p-value <0.05 (Stratagene, CA) and the following cycling conditions: 95° and q-value <0.05, were considered statistically significant. C. for 10 min followed by 40 cycles of 95°C. for 30 sec, 55° The 16S rRNA sequences of significant taxa were used to C. for 1 min and 72°C. for 30 sec. The data acquisition step construct a neighbor-joining with nearest-neighbor inter was set at 55° C. Corynebacteriumtuberculostearicum abun US 2016/0310546 A1 Oct. 27, 2016 10 dance was determined by Q-PCR using a pair of primers B. Example 1 designed in this study: CT-F: 5'-GAACGGAAAGGCCCT GCTTGCA-3 and CTR 5'-GGCTCCTATCCGGTAT Comparative Sinus Microbiota Analysis TAGACC-3'; Lactobacillus sakei abundance was deter mined using the primer pair: LS-F: 5'-GGTAAAGGCT CACCAAG ACCGTGAT-3 and LS-R: 5'TCACGCG (0099 Maxillary sinus samples from 20 subjects (10 CRS GCGTTGCTCCATC-3'. Reaction mixtures (25 ul total) and 10 healthy individuals) were used for this study. Patient contained 10 ng of total DNA, 12.5 ul of 2x QuantiTec details are provided in Table 1. Mucin hyper-secretion is a SYBR green (Qiagen, CA), 2.5ul of each primer and 6.5 ul hallmark of sinus disease (12, 13). Therefore, to confirm that of H.O. Reactions were amplified on the MX3000P Real the CRS patients exhibited a phenotype consistent with Time PCR System (Stratagene, CA) under the following disease, Q-PCR analysis of Muc5A gene (encoding mucin steps: 95°C. for 10 min followed by 40 cycles of 95°C. for primarily secreted from surface epithelium goblet cells in 30 sec, 55° C. for 1 min and 72° C. for 30 sec. The data humans (14, 15)) expression was performed. Muc5A was acquisition step was set at 55° C. significantly up-regulated in CRS patients compared to 0098. For Muc5A expression analysis, confirmed DNA healthy control subjects (FIG. 1), thus validating the pres free total RNA (1 lug) was reverse transcribed at 42°C. for ence of sinus disease in our CRS cohort. Q-PCR analysis of 50 min in a 20 ul reaction mixture (0.025 oligo-dT, 0.5 bacterial burden by total 16S rRNA copy number demon mMdNTPs, 5 ul First Strand buffer, 5 mM MgCl2, 0.01 M strated that peri-operative CRS patients and healthy subjects DTT, and RNAse Out) using 1 ul Superscript II (Invitrogen, exhibited no significant difference in sinus bacterial burden CA). cDNA was diluted 1:5 in molecular grade water. (2.19x10-1.09x10 vs 2.86x10+2.34x10 copies of 16S Reactions were performed in 96 well plates in a 25ul final rRNA gene perug of total DNA in CRS and healthy control volume containing 12.5ul of SYBR Green PCR master mix subjects respectively; ps0.53). The result indicates that the (Qiagen, CA), 2.5 ul of each forward and reverse primer sinus niche can Support a defined bacterial load, and that (final concentration 10 uM per primer; Muc5AF 5'-TGTG microbiota composition and relative taxonomic distribution, GCGGGAAAGACAGC-3' Muc5AR 5'-CCTTCCTATG rather than absolute number of bacteria present, are related GCTTAGCTTCAGC-3'; B-actinF 5'-CACCACACCTTC to disease state. TABLE 1.

Patient information.

Antimicrobial treatment Group Study ID Gender Age SNOT-20 Pre-operative Peri-operative CRS CRS-OO1 M S4 2.75 None Cefazolin CRS CRS-OO2 M S4 2.35 TMPSMX Vancomycin CRS CRS-003 M 48 2.65 None AMPSUL CRS CRS-004 M 33 3.6O None Cefazolin CRS CRS-005 M 56 2.65 Clarithromycin Cefazolin CRS CRS-OO6 F 41 1.85 None Cefazolin CRS CRS-OO7 M 60 3.00 Clarithromycin Levofloxacin CRS CRS-008 M 53 1.55 Ciprofloxacin Cefazolin CRS CRS-009 F 46 2.35 AMOfCLU Cefazolin Control. CRS-010 M 62 0.90 Levofloxacin Centriaxone CRS CRS-O11 M 42 3.35 AMOfCLU Clindamycin Control. CRS-012 M 43 O.15 None Clindamycin Control. CRS-013 F 73 17O None Cefazolin Control. CRS-014 M 41 2.6O None Clindamycin Control. CRS-O15 M 39 O.OO None Clindamycin Control. CRS-016 F 37 2.25 None Cefazolin Control. CRS-017 F 46 O.10 None Clindamycin Control. CRS-018 M 46 2.15 None Clindamycin Control. CRS-019 F 31 OSO None Clindamycin Control. CRS-020 F 18 O3O None Clindamycin Microbiota profiling was not preformed for these subject samples due to insufficient 16S rRNAamplicon; None, no antibiotics administered; TMPSMX, Trimethoprim. Sulfamethoxazole; AMPSUL, Ampicillin Sulbactam; AMOCLU, Amoxicillinclavulanate,

TAC AATGAGC TGC-3' and B-actinR 0100. A standardized phylogenetic microarray, the 16S 5'-ACACCCTGGATAGCAACGTACATGC-3"), 4 ul of rRNAPhyloChip was used to profile the presence and rela diluted cDNA, and 6 ul of molecular grade water. Reactions tive abundance of approximately 8,500 bacterial taxa, rep were amplified on a MX3000P Real-Time PCR System resenting broad membership of all known bacterial phyla under the following conditions: 94° C. for 10 min followed (Hugenholtz phylogenetic classification). This approach was by 40 cycles of 94° C. for 30 sec, 58° C. for 1 min and 72° used in favor of more traditional sequencing approaches to C. for 30 sec. Students t-test was used to calculate p-values maximize the depth of community coverage. Low abun for individual treatment groups, p<0.05 was considered dance microbiome members can contribute considerably to significant. microbiome function (16) and act as keystone species that US 2016/0310546 A1 Oct. 27, 2016 11 shape microbial community composition (17). Using this 0105 Following correction for false discovery (ps0.05, tool, we profiled bacterial communities present in 14 sub qs0.05), a total of 1,482 taxa were detected in significantly jects (7 healthy, 7 CRS) with sufficient amplified 16S rRNA lower relative abundance in CRS patient sinuses, underscor product to be analyzed. Comparative analyses of gross ing the extent of sinus microbiota collapse in the CRS bacterial community metrics between the CRS and healthy patient population. A large number of taxa exhibiting the groups demonstrated that compared to healthy individuals, most significant reductions in relative abundance in the CRS CRS patients exhibited substantial microbiota perturbation, patients belonged to the order Lactobacillales and included characterized by significantly reduced bacterial richness probiotic species such as Lactobacillus sakei, as well as (ps0.005; number of bacterial types detected: FIG. 2A), other phylogenetically distinct lactic acid bacteria (LAB) evenness (ps 0.04; relative distribution of bacterial types: Such as Carnobacterium alterfinditum, Enterococcus FIG. 2B) and diversity (ps0.01; metric calculated using mundtii and Pediococcus pentosaceus implicating LAB in richness and evenness indices; FIG. 2C). maintenance of healthy sinus mucosa. In stark comparison, 0101 Reduced diversity was further demonstrated by only Corynebacteria, in particular, Corynebacterium tuber hierarchical cluster analysis which revealed that the majority culostearicum, (Taxon ID 1493, str. CIP 102346) exhibited a of healthy Subjects clustered in a single tightly-knit group, significant increase in abundance in CRS patients (Correla with bacterial communities compositionally distinct from tion value: 0.6220027; ps0.03, qs0.003). Corynebacterium CRS patients who clustered into two distinct groups (FIG. segmentosum (Taxon ID 1192, str. CIP107068 2D). Healthy subject CRS 14, clustered with CRS patients, (CCUG37878)) was also correlated with CRS (Correlation and it was subsequently determined that this individual had value: 0.5514262; ps0.05). historically suffered from chronic nasal allergies. The pres 0106 The microbiota data were examined to identify ent analysis thus confirms that the sinus microbiota compo those species that correlated with SNOT-20 symptom sever sition of healthy subjects is distinct from that of CRS ity scores, in order to further confirm the clinical signifi patients and, that within the CRS population, discrete sub cance of these findings. A large group of 228 taxa were groups with distinct microbial community profiles exist. significantly (p<0.05) correlated with lower SNOT-20 0102 Antibiotic use was examined, as it influences scores (indicative of healthy sinuses: FIG. 3; Table X). microbiota composition. All study subjects (CRS and Amongst these taxa were members of the LAB, e.g., Lac healthy) received prophylactic antibiotics immediately (1 tobaciliaceae, Enterococcaceae, Aerococcaceae, and Strep hour) prior to Surgery and sample collection according to tococcaceae, further Supporting the idea that these families standard practice. Long-term pre-operative antibiotic use are protective in healthy sinuses. was absent for the majority of healthy subjects, and variable 0107 As noted above, the relative abundance of only two in the CRS patient group (Table 1). Patients with disparate bacterial species was positively correlated with increased long-term antimicrobial administration histories (e.g. CRS symptom severity; both belonged to the Corynebacteriaceae, 4, 6 and 2: Group II: Table 1) clustered closely together. with C. tuberculostearicum most correlated with symptom Moreover, Group II included two patients who had not severity (r=0.62; ps0.02). Q-PCR analysis was performed received long-term antibiotics, and who exhibited signifi using primers designed to specifically amplify C. tubercu cantly lower (p<0.006) sinus community diversity compared lostearicum to validate these findings. Linear regression of to microbiota from patients in Group I. Group I subjects had C. tuberculostearicum Q-PCR-derived copy number against received long-term prophylactic antibiotic administration, both array-reported fluorescence intensity or against SNOT indicating that while antibiotic therapy may contribute, it is 20 score demonstrated concordance (r=0.66; ps0.01 and not the sole selective pressure defining bacterial community r=0.68; ps0.01 respectively). The regression data thus cor composition and loss of diversity in this niche. roborate the array-based findings and confirm a strong relationship between the abundance of this species and sinus C. Example 2 symptom severity (FIGS. 4A-4B). Taxa Characteristic of Healthy and CRS Sinuses D. Example 3 0103 Results from the Sino-Nasal Outcomes Test (SNOT-20) questionnaire, a metric to score sinus sympto Murine Model of Sinusitis mology, confirmed that CRS patients reported significantly 0.108 To determine whether C. tuberculostearicum, higher scores (i.e., more severe sinus symptomology (ps0. which is typically considered a skin commensal, exhibited 003)) than healthy controls. The observed microbiota clus any pathogenic potential, a murine model of sinus infection tering patterns were consistent with patient-reported sinus was developed. Goblet cell hyperplasia and mucin hyperse symptomology, as shown in FIG. 2D). Given this level of cretion were used as indicators of pathology (12), and as the independent validation of disease activity, specific commu outcome measures to define pathogenic activity in the nity members that differentiated healthy subjects and CRS sinuses. This model is also allows for determination of patients were next characterized at the taxonomic level. whether C. tuberculostearicum is influenced by resident 0104 Known pathogenic members of the microbiota in the sinus cavity. Pseudomonadaceae, Lachnospiraceae, Ralstoniaceae, 0109 Four groups of mice (n=5 animals per group) Mycobacteriaceae and Helicobacteriaceae were detected in representing: i. Untreated control; ii. Antibiotic treated (to both CRS patients with, and healthy subjects without, elicit microbiome depletion); iii. C. tuberculostearicum sinonasal symptoms. Thus, the mere detection of a Suspected (ATCC #35694) inoculated; and iv. Antibiotic treated and C. or known pathogen in a given niche does not necessarily tuberculostearicum inoculated animals were used (FIG. 5). imply pathogenic activity, indicating that the microbiota Q-PCR analyses of total 16S rRNA copy number from the composition at a given site may play a large role in defining sinuses of these animals confirmed that the burden of the activity of community members. bacteria in the antibiotic treated groups was significantly US 2016/0310546 A1 Oct. 27, 2016

lower (p-0.03) than that of untreated animals. The data the presence of these species (FIG. 7). Histological imaging confirm acute antimicrobial depletion of bacterial burden of the maxillary sinuses (FIG. 8A) demonstrated again that and mucosal sinus microbiota diversity in antibiotic-treated the antibiotic-treated and C. tuberculostearicum inoculated groups. Histological examination of the sinus mucosa from group exhibited significant increases in goblet cell hyper each group demonstrated that the untreated control and plasia and mucin hypersecretion (FIG. 8A, panel iii). Mice antibiotic treated animals did not exhibit aberrant epithelial receiving identical numbers of L. Sakei, however, demon physiology (FIG. 6A, panels i and ii), while instillation of C. strated epithelial physiology comparable to that of control tuberculostearicum in the presence of a replete sinus micro animals (no significant differences in goblet cell numbers; Table 3: FIG. 8A, panel iv). That L. sakei inoculated animals biota elicited a modest increase in the number of mucin did not display the pathogenic response indicated that it was secreting goblet cells (FIG. 6A, panel iii). specifically due to C. tuberculostearium. Enumeration of 0110 Animals treated with both antimicrobial and C. goblet cell numbers in each treatment group confirmed these tuberculostearicum exhibited profound goblet cell hyperpla observations (FIG. 8B: Table 3). sia, (FIG. 6A, panel iv: FIG. 6B and Table 2), significantly 0113. The collective data indicate that L. sakei is protec greater than that observed in any other group. Since mucin tive in the sinus mucosa. We next investigated whether hypersecretion is a hallmark of respiratory infection (18) and co-instillation of L. Sakei with C. tuberculostearicum could chronic sinusitis (12, 13), these data confirm that C. tuber abrogate the goblet cell hyperplasia and mucin hypersecre culostearicum is capable of inducing a characteristic tion phenotype induced by C. tuberculostearicum, even in response to pathogenic microbes and that this response is the context of a depleted native microbiota. Following significantly augmented under conditions of depleted sinus treatment with antibiotics, equal numbers of both species microbiota. were instilled into the sinuses of mice. Histological exami nation revealed sinus epithelia comparable to that of animals TABLE 2 in the control groups (FIG. 8A, panel V), with no significant p-values for goblet cell number per Lm of epithelial cell Surface differences in goblet cell numbers observed across these compared across mouse treatment groups (FIGS. 6A-6B). groups (FIG. 8B and Table 3). Q-PCR analyses demon strated significantly (p<0.02) reduced C. tuberculostearicum AB- CT- AB+ CT- AB- CT AB+CT+ abundance in the co-instilled animals compared to animals AB- CT- O.36 O.O11 O.OO34 infected with C. tuberculostearicum alone (FIG. 7). L. sakei AB+ CT- O.OO12 O.OOO1 numbers in these animals were, however, similar to those in AB- CT 0.0076 animals treated with L. sakei alone (FIGS. 8A-8B). The data AB+CT+ indicate that L. Sakei protects the sinus epithelium, puta tively through competitive inhibition of C. tuberculosteari 0111. The above experiment was repeated, with inclusion cum. L. Sakei thus represents a novel, probiotic therapy for of an additional group treated with antibiotics prior to amelioration or prevention of sinus pathology, even in instillation of L. sakei (ATCC 15521) (FIG. 5), to demon patients with severe sinus microbiome depletion. strate that goblet cell hyperplasia and mucin hypersecretion were induced specifically by C. tuberculostearicum. The TABLE 3 conditions were designed to confirm that the characteristic Significance values for comparative goblet cell number per Lim of pathogenic response did not simply represent a host epithelial cell surface across mouse treatment groups (FIGS. 8A-8B). response to instillation of any bacterial species into the sinus niche. L. Sakei was selected because this species was present AB+ AB+CT+ in high abundance in healthy mucosal samples, and was the AB- CT- CT- AB+ LS+ AB+CT+ LS+ most significantly- depleted taxon in CRS patients. These AB- CT- 1 O.2002 0.0099 0.2082 results indicated that L. Sakei is a protective sinus mucosal AB+ CT- O.2002 0.0099 0.2082 colonizer, and likely would not induce a pathogenic AB+ LS+ O.O118 O.S717 SOS AB+CT+ O.O141 ponse. AB+CT+ LS+ 0112 Q-PCR was performed on all treatment groups to confirm that animals receiving bacterial inocula exhibited TABLE X Bacterial species negatively correlated with sinus sympton severity Phylum Family Taxon prokMSA Name Estimate P value Q value Firmicutes Lactobacillaceae 3547 Lactobacilius frumenti str. -O.S324S16 O.04998019 0.035270589 TMW 1.666 Proteobacteria Helicobacteraceae 10520 Helicobacter sp. blood -O.S32S229 O.O4994373 0.035270589 isolate 964 Proteobacteria Campylobacteraceae 10538 Arcobacter cryaerophilus -O.S332394 O.O495.7863 0.035270589 Unclassified Unclassified 243 hot spring clone OPB25 -O.S332699 O.O4956314 0.035270589 Proteobacteria Nitrospinaceae 594 uranium mining mill tailing -O.S334925 O.O4945O17 0.035270589 clone GR-296.II.52 GR 296.I.S2 Firmicutes Acholeplasmataceae 3977 Chinaberry yellows -0.53364 O.O493.7536 0.035270589 phytoplasma str. CbY1 US 2016/0310546 A1 Oct. 27, 2016 13

TABLE X-continued Bacterial species negatively correlated with sinus Sympton severity Phylum Family Taxon prokMSA Name Estimate P value Q value Bacteroidetes Porphyromonadaceae S429 -O.S336SS8 O.O4936,737 O.O35270589 Cyanobacteria Chloroplasts 5040 Solantin nigrum -O.S336963 O.04934686 O.O35270589 Proteobacteria Bradyrhizobiaceae 7390 Afipia genosp. 10 str. G8996 -O.S336971 O.04934647 O.O35270589 Firmicutes Lachnospiraceae 2994 termite gut clone RS-L15 -O.S33732 O.O493.2878 O.O35270589 Synergistes Unclassified termite gut homogenate -0.5342366 O.O4907378 O.O35270589 clone RS-D89 Bacteroidetes Flavobacteriaceae Capnocytophaga gingivais -0.5343746 O.04900422 O.O35270589 Str. ChDC OS45 Proteobacteria Desulfobacteraceae 10046 Desulfobacterium cetonicum -O.S343791 O.O35270589 str. DSM 7267 oil recovery Water Verrucomicrobia Verrucomicrobiaceae 613 hydrothermal vent -0.5344217 O.04898.049 O.O35270589 polychaete mucous clone P. palm C 85 Proteobacteria Unclassified 10427 hydrothermal vent 9 degrees -0.5344767 O.04895279 O.O35270589 North East Rise Pacific Ocean clone CH3 17 BAC 16SrRNA 9N EPR Actinobacteria Micrococcaceae 1686 Yania haiotoierans str. YIM -O.S345.007 O.04894071 O.O35270589 70085 Unclassified Onclassified 4.410 O.0489.1381 O.O35270589 Cyanobacteria Onclassified SO10 LPP-group cyanobacterium -0.5346143 O.048.88353 O.O35270589 isolate str. QSSC5cya QSSC5cya Acidobacteria Onclassified 6357 anaerobic benzene-degrading -O.S346545 O.0488.6331 O.O35270589 clone Cart-N4 Proteobacteria Onclassified 8961 Calyptogena magnifica O.04871181 O.O35270589 symbiont Firmicutes Acholeplasmataceae 3945 Ash witches-broom -O.S352908 O.04854413 O.O35270589 phytoplasma str. Ash WB Firmicutes Lachnospiraceae 2693 ruminantium str. GA195 -0.53S3908 O.048494.09 O.O35270589 Firmicutes Peptococci 865 Selenomonas splitigena str. -O.S354898 O.0484.4462 O.O35270589 Acidaminococc ATCC 35.185 Firmicutes Paenibacillaceae 3641 Brevibacilius sp. MN 47.2a -0.5355341 O.0484224.5 O.O35270589 Planctomycetes Planctomycetaceae 48SS aerobic basin clone -0.5.360602 O.04816O2 O.O35270589 CYOARAO26D10 Firmicutes Lactobacillaceae 3696 Lactobacilius kalixensis str. -O.S36599 O.O.4789264 O.O35270589 Kx127A2: LMG 22115T: DSM 16043T; CCUG 48459T Proteobacteria Onclassified 10259 -O.S369322 0.0477 2772 O.O35270589 Firmicutes Onclassified 2541 UASB granular sludge clone -O.S369672 O.0477104.4 O.O35270589 UT2 Firmicutes Lachnospiraceae 2681 termite gut homogenate -0.5370585 O.04766533 O.O35270589 clone RS-K41 bacterium Proteobacteria Bradyrhizobiaceae 7126 ground water deep-well -O.S371332 O.04762847 O.O35270589 injection disposal site radioactive wastes Tomsk-7 clone S15A-MN96 proteobacterium Firmicutes Syntrophomonadaceae 2483 trichloroethene-contaminated -O.S376491 O.O4737434 O.O35270589 site clone FTLM142 bacterium Firmicutes Peptococci 131 pig feces clone -O.S377144 O.O4734226 O.O35270589 Acidaminococc Proteobacteria Onclassified 8587 Mars Odyssey Orbiter and -0.5377735 O.O4731322 O.O35270589 encapsulation facility clone TS-3 Bacteroidetes Flavobacteriaceae S906 Capnocytophaga Spittigena -O.S378696 O.O35270589 Str. ATCC 33612 Bacteroidetes Flavobacteriaceae 5933 Flavobacterium columnare -O.S378938 O.O35270589 str. PH-97028 (IAM 14821) Firmicutes Lachnospiraceae 296S oral endodontic infection -O.S381045 O.O35270589 clone MCE9 173 Actinobacteria Micrococcaceae 1494 Arthrobacter agilis str. DSM -O.S383154 0.04704757 O.O35270589 2O550 BRC1 Onclassified 5143 soil clone PBS-II-1 -O.S383959 O.O35270589 Firmicutes Enterococcaceae 3.298 Enterococci is saccharolyticus -O.S391099 O.O35270589 Str. LMG 11427 Cyanobacteria Onclassified SO38 Rumen isolate str. YS2 -053928.66 O.O4657427 O.O35270589 Firmicutes Lachnospiraceae 433S termite gut homogenate -O.S392971 O.O4656915 O.O35270589 clone RS-N86 bacterium US 2016/0310546 A1 Oct. 27, 2016 14

TABLE X-continued Bacterial species negatively correlated with sinus Sympton severity Phylum Family Taxon prokMSA Name Estimate P value Q value Firmicutes Aerococcaceae Mariniiactibacilius -O.S397087 O.O4636966 O.O35270589 psychrotolerans str. O21 Bacteroidetes Unclassified 5257 marine? clone KD3-67 -O.S398265 O.O35270589 Firmicutes Peptococci 534 chlorobenzene-degrading -O-5401839 O.O35270589 Acidaminococc consortium clone IIA-26 Spirochaetes Spirochaetaceae 6526 Treponema sp. str. 7CPL208 -O-5402284 O.O461186 O.O35270589 Actinobacteria Micrococcaceae 1573 Arthrobacier nicotianae str. -O-5405694 O.04595441 O.O35270589 SB42 Proteobacteria Desulfoarculaceae 10227 marine sediment clone Bol11 -O.S4O9671 O.04576348 O.O35270589 Firmicutes Clostridiaceae 4278 granular sludge clone R1 p16 -O.S4O9777 O.04575841 O.O35270589 Proteobacteria Unclassified 7060 -O.S41O3S3 O.04573O8 O.O35270589 Bacteroidetes Unclassified 5785 Mono Lake at depth 35 m -0.5413643 O.O35270589 station 6 Jul. 2000 clone ML635J-56 Firmicutes Streptococcaceae 3499 Streptococci is constellatus -0.541.6222 O.O35270589 Str. ATCC27823 Firmicutes Clostridiaceae 4177 Cliostridium subterminale -0.5419209 O.O35270589 DSM 2636 Bacteroidetes Rikenellaceae S889 termite gut homogenate -0.542O021 O.O452693 O.O35270589 clone RS-F73 bacterium Proteobacteria Bdellowibrionaceae 10010 uranium mining waste pile -0.5422324 O.O35270589 clone JG37-AG-139 proteobacterium Firmicutes Peptococci 392 oral endodontic infection -O-5424949 O.O4SO3S37 O.O35270589 Acidaminococc clone MCE7 134 Bacteroidetes Sphingobacteriaceae 5513 crevicular epithelial cells -O.S425299 O.O4SO1877 O.O35270589 clone AZ123 Firmicutes Mycoplasmataceae 3929 Mycoplasma -0.5427O64 O.O4493525 O.O35270589 gypsbengalensis str. Gb-V33 Firmicutes Clostridiaceae 43OO termite gut clone RS-060 -0.5428522 O.04486631 O.O35270589 Proteobacteria Campylobacteraceae 10484 Campylobacter helveticus -O.S430959 O.O447S126 O.O35270589 OP10 Unclassified S1.4 sludge clone SBRA136 -O.S432395 O.O4468356 O.O35270589 Proteobacteria Syntrophobacteraceae 10221 granular sludge clone R3p4 -O.S433823 O.O4461634 O.O35270589 Firmicutes Lactobacillaceae 3330 Lactobacilius kitasatonis str. -O.S437728 O.04443286 O.O35270589 KM9212 BRC1 Unclassified 118 penguin droppings sediments -0.544O147 O.04431948 O.O35270589 clone KD1-1 Firmicutes Peptococci 39 forested wetland clone -0.5441 698 O.04424689 O.O35270589 Acidaminococc RCP2-71 Proteobacteria Pasteurellaceae 92.13 Haemophilus quientini str. -0.544316 O.O44-17854 O.O35270589 MCCM 02026 Firmicutes Streptococcaceae 3629 Streptococci is maitans str. -0.5443712 O.O4415277 O.O35270589 UA96 Proteobacteria Unclassified 9760 deep marine sediment clone -O.S4SO115 O.0438545 O.O35270589 MB-A2-137 Firmicutes Unclassified 926 -O.S457.011 O.043S3492 O.O35270589 Firmicutes Staphylococcaceae 3524 Gemella haemolysans -O.S458225 O.04347884 O.O35270589 Proteobacteria Syntrophobacteraceae 10O21 uranium mill tailings soil -O.S46OSS O.04337153 O.O35270589 sample clone Sh765B-TZT 29 proteobacterium Actinobacteria Micrococcaceae 1593 Arthrobacter globiformis -O.S462S32 O.O35270589 Proteobacteria Desulfobacteraceae 9875 hydrothermal sediment clone -0.54634.08 O.04323997 O.O35270589 AF42O3S4 Cyanobacteria Chloroplasts 4966 Adiantum pedatin -0.547O294 O.04292415 O.O35270589 Bacteroidetes Prevotellaceae 5893 tongue dorsa clone DOO45 -O.S471467 O.O428 7049 O.O35270589 Firmicutes Peptococci Acidaminococc oral endodontic infection -O.S472263 O.04283412 O.O35270589 clone MCE10 265 Chlorobi Unclassified 636 benzene-degrading nitrate- -O.S473972 O.O4275611 O.O35270589 reducing consortium clone Cart-N3 bacterium Firmicutes Carnobacteriaceae 3536 -0.5475614 O.O42681.28 O.O35270589 Bacteroidetes Flavobacteriaceae 5473 -0.5478.064 O.O4256979 O.O35270589 Proteobacteria Syntrophobacteraceae 9845 uranium mining waste pile -O.S4.84553 O.O35270589 clone JG37-AG-128 proteobacterium Proteobacteria Campylobacteraceae 10540 Campylobacter Showae str. -0.5489,328 O.O420599 O.O35270589 LMG 12636 Proteobacteria Sphingomonadaceae 7100 Novosphingobium -O.S490852 O.04.1991.26 O.O35270589 capsulatum str. GIFU11526 Proteobacteria Bradyrhizobiaceae 6887 Bradyrhizobium str. YB2 -0.5492974 O.O35270589 Chloroflexi Unclassified 76 DCP-dechlorinating -O.SSOO398 O.O35270589 consortium clone SHA-147 US 2016/0310546 A1 Oct. 27, 2016 15

TABLE X-continued Bacterial species negatively correlated with sinus Sympton severity Phylum Family Taxon prokMSA Name Estimate P value Q value Firmicutes Clostridiaceae 426S termite gut homogenate -O.SSOOS63 O.04 55579 O.O35270589 clone RS-N70 bacterium Bacteroidetes Flavobacteriaceae 5436 Arctic sea ice ARK10004 -0.55O1646 O.04 SO741 O.O35270589 Actinobacteria Unclassified 1898 termite gut homogenate -O.SSO3S48 O.04 4226 O.O35270589 clone RS-J10 bacterium Firmicutes Aerococcaceae 3631 Abiotrophia defectiva str. -O.SSO4S43 O.04 37826 O.O35270589 GIFU12707 (ATCC49176) Firmicutes Streptococcaceae 3753 Streptococcus suis str. 8074 -O.SSOS636 O.04 32961 O.O35270589 Bacteroidetes Flexibacteraceae 6124 Flexibacter flexiiis subsp. -0.5507177 O.04 26111 O.O35270589 pelliculosus str. IFO 16028 Subsp. Firmicutes Enterococcaceae 3382 -O.SSO8457 O.04 20423 O.O35270589 Firmicutes Leuconostocaceae 3573 Leticonostoc fictinenim str. -O.SS11595 O.04 O.O35270589 FS-1 Firmicutes Streptococcaceae 3422 Streptococcus thermophilus -O.SS11839 O.04 OS433 O.O35270589 Str. DSM 20617 Firmicutes Lachnospiraceae 4315 termite gut homogenate -O.SS12027 O.04 O4598 O.O35270589 clone RS-N94 bacterium Onclassified Unclassified 7444 -O.SS13268 O.O4O991 O7 O.O35270589 Bacteroidetes Flexibacteraceae 6261 Arctic Sea ice cryoconite -0.5515557 O.04.088996 O.O35270589 clone ARKCRY-50 Planctomycetes Planctomycetaceae 4948 anoxic basin clone -0.5S24994 O.O404749 O.O35270589 CYOARAO27DO1 Firmicutes Streptococcaceae 3253 derived cheese sample clone -0.5527952 O.04034546 O.O35270589 32CR Firmicutes Clostridiaceae 4614 Clostridium sp. str. JC3 -O.SS342S1 O.04007081 O.O35270589 Bacteroidetes Flavobacteriaceae 5267 bacterioplankton clone -O.SS35169 O.O4003091 O.O35270589 AEGEAN 179 Natronoanaerobium Onclassified 4377 Mono Lake at depth 35 m -0.5538.773 O.O39874.48 O.O35270589 station 6 Jul. 2000 clone ML635J-6S G - C Onclassified 2489 Guaymas Basin -O.SS43007 O.O396.913 O.O35270589 hydrothermal vent sediments clone BO1R005 Chloroflexi Onclassified 2534 orest soil clone S085 -0.5543422 O.O3967338 O.O35270589 Bacteroidetes Onclassified 5957 hydrothermal vent -O.SS43543 O.O396.6819 O.O35270589 polychaete mucous clone P. palm CA 20 Firmicutes Lactobacillaceae 3521 Pediococci is inopiniatus str. -O.SSS167 O.O3931844 O.O35270589 DSM 20285 Firmicutes Onclassified 77 hermal soil clone YNPFFP9 -O.SSS2835 O.O3926852 O.O35270589 Firmicutes Clostridiaceae 4550 Swine intestine clone p-320- -O.SSS8022 O.O390467 O.O35270589 a3 Firmicutes Lachnospiraceae 2668 ermite gut homogenate -O.SSS8071 O.O3904461 O.O35270589 clone RS-G40 bacterium Actinobacteria Micrococcaceae 1557 Arthrobacter oxydans str. -O.SS60342 O.O3894779 O.O35270589 DSM 2011.9 Firmicutes Clostridiaceae 4459 ermite gut clone RS-109 -O.S.S.67823 O.O3863011 O.O35270589 Bacteroidetes Flavobacteriaceae 5521 Flavobacteriaceae str. -O.SS68853 O.O38586SS O.O35270589 SW269 Proteobacteria Campylobacteraceae 10397 groundwater clone 1006 -O.SS70026 O.O35270589 Proteobacteria Onclassified 9876 deep marine sediment clone -O.SS7006S O.O35270589 MB-B2-106 Firmicutes Lachnospiraceae 4514 termite gut homogenate -O.SS7O651 O.O3851051 O.O35270589 clone RS-B34 bacterium Proteobacteria C Olyangliaceae 9733 bacterioplankton clone -O.SS74153 O.O3836281 O.O35270589 ZA373Sc Firmicutes Onclassified 522 UASB granular sludge clone -0.5575107 O.O3832264 O.O35270589 UT1 Firmicutes Streptococcaceae 3722 Lactococcus II 1403 subsp. -O.SS78807 O.O381 6718 O.O35270589 iaciis str. IL1403 Firmicutes Peptococci Selenomonas ruminantium -0.5580707 O.O35270589 Acidaminococc Str.JCM6582 Firmicutes Lactobacillaceae 3885 Pediococci is peritosaceus -O.SS82O24 O.O35270589 Proteobacteria Sphingomonadaceae 7440 Sphingobium chungbukense -O.SS8408 O.O35270589 Str. DJ77 Firmicutes Aerococcaceae 3326 Nostocoida innicola I str. -O.SS8438 O.O3793384 O.O35270589 Ben2O6 Bacteroidetes Flexibacteraceae S602 -O.SS8608 O.O3786285 O.O35270589 Proteobacteria Polyangiaceae 10298 marine tidal mat clone -O.SS88222 0.03777357 O.O35270589 BTM36 Firmicutes Bacillaceae 3900 Bacilius licheniformis str. -O.SS883.43 0.03776853 O.O35270589 DSM 13 US 2016/0310546 A1 Oct. 27, 2016 16

TABLE X-continued Bacterial species negatively correlated with sinus Sympton severity Phylum Family Taxon prokMSA Name Estimate P value Q value Bacteroidetes Rikenellaceae S892 anoxic bulk soil flooded rice -O.S591.33 O.O3764434 O.O35270589 microcosm clone BSV73 Proteobacteria Pasteurellaceae 8195 Haemophilus influenzae Str. -O.S.S91662 O.O3763OS3 O.O35270589 R2866 Chlorobi Chlorobiaceae 859 Chlorobium phaeovibrioides -O.S.S98611 O.O3734276 O.O35270589 Str. 2631 Firmicutes Clostridiaceae 4477 termite gut homogenate -0.5599449 O.O373O819 O.O35270589 clone RS-N85 bacterium Proteobacteria Onclassified 244 deep marine sediment clone -O.S600791 O.O372S282 O.O35270589 MB-C2-152 Bacteroidetes Flavobacteriaceae 6248 Capnocytophaga sp. oral -0.56O46 O.O37096O7 O.O35270589 strain str. S3 Proteobacteria Onclassified acid mine drainage clone -O-5606514 O.O37O1749 O.O35270589 AS6 Firmicutes Streptococcaceae 3588 Streptococci is downei Str. -O.S606SS6 0.03701576 O.O35270589 ATCC 33748 Firmicutes Peptococci 1036 Great Artesian Basin clone -O.S60657 O.O3701S2 O.O35270589 Acidaminococc GO7 Chloroflexi Onclassified 2397 deep marine sediment clone -0.561111 O.O368,293 O.O35270589 MB-C2-27 Actinobacteria Micrococcaceae 2020 Rothia deniocariosa str. -O.S612027 O.036791.83 O.O35270589 CDC B2OO Bacteroidetes Onclassified 5353 trichloroethene-contaminated -O.S6145S O.O366.8889 O.O35270589 site clone FTLpost3 bacterium Bacteroidetes Flavobacteriaceae 5991 Tenacibaculum ovolyticum -O.S617321 O.O36576OS O.O35270589 Str. IAM14318 Proteobacteria Sphingomonadaceae 7036 Litibacterium antiloederans -O.S62OSS O.O3644492 O.O35270589 Str. LC8 Cyanobacteria Chloroplasts 5147 Emiliania huxleyi str. -O.S62O697 O.O3643899 O.O35270589 Plymouth Marine Laborator PML 92 Actinobacteria Acidimicrobiaceae 1090 -O.S623333 O.03633219 O.O35270589 Proteobacteria Unclassified 98.13 hydrothermal sediment clone -0.5624152 O.O36299.06 O.O35270589 AF42O340 Firmicutes Enterococcaceae 3433 Tetragenococci is matriaticits -0.5626691 O.O361.96S1 O.O35270589 Firmicutes Acholeplasmataceae 4044 -O.S626807 O.O3619181 O.O35270589 TM7 Unclassified 8040 oral cavity clone BE109 -O.S630985 O.O36O23S4 O.O35270589 Bacteroidetes Porphyromonadaceae 5295 Swine intestine clone p-987- -0.563282 O.O3S94978 O.O35270589 S962-5 Bacteroidetes Flavobacteriaceae S914 Psychroserpens burtonensis -O.S63714 0.03577667 O.O35270589 str. S2-64 Firmicutes Lachnospiraceae 4613 rumen clone 3COd-3 -O.S638647 O.O3571638 O.O35270589 Firmicutes Acholeplasmataceae 4046 Pigeon pea witches”-broom -0.56396 O.O3567834 O.O35270589 mycoplasma-like organism Bacteroidetes Flavobacteriaceae 6269 acidic forest soil clone UC1 -0.564O842 O.O3S62876 O.O35270589 OP9 JS1 Unclassified 969 DCP-dechlorinating -0.5641236 O.03561305 O.O35270589 consortium clone SHA-1 Firmicutes Clostridiaceae 418O termite gut homogenate -0.5642O19 0.035581.87 O.O35270589 clone RS-M23 bacterium Cyanobacteria Chloroplasts 4976 Calypogeia mieileriana -0.5642267 0.035571.99 O.O35270589 Firmicutes Enterococcaceae 3261 Enterococcus mindtii str. -0.5647034 O.O3S38246 O.O35270589 LMG 10748 Actinobacteria Micrococcaceae 1324 glacial ice isolate str. -O.S6S5964 O.O3SO2939 O.O35270589 CanDirty1 Firmicutes Peptococci 761 Anaeroglobits geminati is str. -O.S657127 O.O349836 O.O35270589 Acidaminococc AIP313.00; CIP 106856; CCUG 44773 Bacteroidetes Onclassified 5475 SHA-25 clone -O.S66O295 O.O3485913 O.O35270589 Planctomycetes Planctomycetaceae 4831 Planctomyces brasiliensis -O.S66052 O.O348SO3 O.O35270589 Firmicutes Lachnospiraceae 4510 termite gut homogenate -O.S663472 O.O347346 O.O35270589 clone RS-Q53 bacterium Firmicutes Lactobacillaceae 3767 Lactobacilius Stiebicits str. -O.S665992 O.O34636O7 O.O35270589 CECT 5917T Firmicutes Peptococci 709 Selenomonas ruminantium -O.S670817 O.O3444798 O.O35270589 Acidaminococc Str. S2O Proteobacteria Onclassified 98.84 forested wetland clone -0.5671464 O.O3442282 O.O35270589 RCP2-62 Firmicutes LeuconoStocaceae 3497 Weissella koreensis S-5673 -O.S673939 O.O3432668 O.O35270589 Firmicutes Enterococcaceae 3713 Enterococcits Cecorum str. -0.5677953 O.O3417121 O.O35270589 ATCC431.98 Firmicutes Lactobacillaceae 3566 Lactobacilius pontis str. LTH -O.S6907S6 O.O3367873 O.O35270589 2587 US 2016/0310546 A1 Oct. 27, 2016 17

TABLE X-continued Bacterial species negatively correlated with sinus Sympton severity Phylum Family Taxon prokMSA Name Estimate P value Q value Firmicutes Onclassified 4536 Mono Lake at depth 35 m -O.S692171 O.O3362463 O.O35270589 station 6 Jul. 2000 clone ML635J-14 G -- C Firmicutes Onclassified 3481 -0.5692343 O.O3361806 O.O35270589 Proteobacteria Syntrophobacteraceae 9661 DCP-dechlorinating -0.5696648 O.O334539 O.O35270589 consortium clone SHD-1 Proteobacteria Desulfobacteraceae 10364 marine Surface sediment -0.56971S2 O.O3343469 O.O35270589 clone SB2 Firmicutes Lachnospiraceae 2943 human thigh wound isolate -0.5708144 O.O3301837 O.O35270589 Str. MDA2477 Firmicutes Enterococcaceae 3318 Enterococcits ratii str. ATCC -0.571 1715 O.O3288393 O.O35270589 7OO914 Bacteroidetes Porphyromonadaceae 6O12 mouse feces clone L11-6 -0.5712O18 0.03287255 O.O35270589 Firmicutes Clostridiaceae 4554 termite gut clone RS-068 -0.571372 O.O328O865 O.O35270589 Firmicutes Lactobacillaceae 3490 Lactobacilius Suntoryeus str. -O.S71473S 0.03.277059 O.O35270589 LH Firmicutes Clostridiaceae 4582 Swine intestine clone p-2600- -O.S72SO13 O.03238692 O.O35270589 9F5 Firmicutes Peptococci 242 Destifosporosini is orientis -O.S726663 O.O3232S64 O.O35270589 Acidaminococc Str. DSMZ 7493 Firmicutes Streptococcaceae 3907 aortic heart valve patient -0.57 27726 O.O3228622 O.O35270589 with endocarditis clone v6 Firmicutes Peptococci 710 Centipedia periodontii Str. -O.S72926S O.O3222922 O.O35270589 Acidaminococc HB-2 Cyanobacteria Chloroplasts S183 Pisum sativum-chloroplast -O.S756529 O.O31231.33 O.O35270589 Cyanobacteria Chloroplasts 5.192 Cycas revolutta -O.S759699 O.O3111681 O.O35270589 Firmicutes Lachnospiraceae 2991 rumen clone 3C3d-8 -O.S761139 O.O3106487 O.O35270589 Firmicutes Enterococcaceae 3881 Enterococci is dispar str. -O.S764427 O.O3O94657 O.O35270589 LMG 13521 Bacteroidetes Flavobacteriaceae S836 Capnocytophaga granulosa -0.5766249 O.O3O881.16 O.O35270589 str. LMG 12119: FDC SD4 Firmicutes Peptococci 3OO benzene-contaminated -0.5787987 O.O3O10865 O.O35270589 Acidaminococc groundwater clone ZZ12C8 Bacteroidetes Onclassified 5784 fruiting body Pleurotus -O.S792647 O.O2994496 O.O35270589 eryngii clone PEO1 Firmicutes Bacillaceae 3848 Bacillus sp. str. TUT1007 -0.57939S4 O.O298.991S O.O35270589 Firmicutes Carnobacteriaceae 3792 Carnobacterium sp. str. D35 -O-5804968 O.O2951531 O.O35270589 Firmicutes Lachnospiraceae 4533 termite gut homogenate -O.S81153 O.O292.8835 O.O35270589 clone RS-NO6 bacterium Onclassified Onclassified 651 -O.S811737 O.O29281.21 O.O35270589 Bacteroidetes Flavobacteriaceae 6246 crevicular epithelial cells -O.S814157 O.O291.9784 O.O35270589 clone BUO84 Firmicutes Lachnospiraceae 4434 termite gut homogenate -O-5814826 O.O2917485 O.O35270589 clone RS-K11 bacterium Firmicutes Lactobacillaceae 3418 Lactobacilius Subsp. aviaritis -O-5814892 O.O2917259 O.O35270589 Firmicutes Clostridiaceae 4297 -O.S818952 O.O2903323 O.O35270589 Proteobacteria Polyangiaceae 9671 hydrothermal sediment clone -O.S821323 O.028952O6 O.O35270589 AF42O357 Firmicutes Acholeplasmataceae 3976 -0.5828049 O.O287228 O.O35270589 Firmicutes Acholeplasmataceae 3961 Clover yellow edge -O-5834912 O.O284.9025 O.O35270589 mycoplasma-like organism Bacteroidetes Flavobacteriaceae 5301 -0.583.8861 O.O35270589 Firmicutes Streptococcaceae 3685 Streptococci is gordonii Str. -O-584O426 O.O35270589 ATCC 10558 Actinobacteria Micrococcaceae 2063 Rothia deniocariosa str. -O.S849.591 O.O27997 S4 O.O35270589 ATCC 17931 Firmicutes Clostridiaceae 4272 termite gut homogenate -0.5856846 O.O35270589 clone RS-M34 bacterium Synergistes Unclassified 353 UASB reactor granular -O-5862744 0.02756153 O.O35270589 sludge clone PD-UASB-13 G + C Fusobacteria Fusobacteriaceae 721 human mouth clone -O.S8637O6 O.O2752982 O.O35270589 P2PB 51 Firmicutes Clostridiaceae 4524 termite gut clone RS-093 -O.S872.218 O.0272506 O.O35270589 Firmicutes Acholeplasmataceae 4045 Chinaberry yellows -O.S87.4789 O.O271667 O.O35270589 phytoplasma Firmicutes Lachnospiraceae 41SS termite gut homogenate -O.S883278 O.O35270589 clone RS-K92 bacterium Firmicutes Lactobacillaceae 3703 Lactobacilius Saivarius Str. -O.S896OSS O.O2647992 O.O35270589 RA2115 Firmicutes Clostridiaceae 4622 termite gut clone RS-L36 -0.590871 O.O35270589 Firmicutes Aerococcaceae 3833 Carnobacterium -0.5911531 O.O35270589 alterfunditum US 2016/0310546 A1 Oct. 27, 2016 18

TABLE X-continued Bacterial species negatively correlated with sinus Sympton severity Phylum Family Taxon prokMSA Name Estimate P value Q value Firmicutes Lachnospiraceae 4540 termite gut homogenate -0.5918.608 O.O2S76588 O.O35270589 clone RS-M18 bacterium Firmicutes Streptococcaceae 3251 Streptococcus cristatus str. -0.59192O6 O.O2S74714 O.O35270589 ATCC S1100 TM7 Unclassified 2697 midgut homogenate -O.S91935 O.O2S7426S O.O35270589 Pachnoda ephippiata larva clone PeM47 Chloroflexi Unclassified 258 DCP-dechlorinating -0.5925901 O.O2SS3809 O.O35270589 consortium clone SHD-14 Firmicutes Aerococcaceae 3840 Trichococcus pasteurii str. -0.5926.609 O.O2SS1607 O.O35270589 KoTa2 Proteobacteria Neisseriaceae 8143 subgingival dental plaque -O.S935762 O.O2S232S3 O.O35270589 clone AK105 Firmicutes Acholeplasmataceae 3975 Black raspberry witches”- -0.594OO16 O.O2S1 O154 O.O35270589 broom phytoplasma str. BRWB witches”-broom Firmicutes Streptococcaceae 3543 -0.5975365 O.O2403278 O.O35270589 Chloroflexi Onclassified 2339 uranium mill tailings soil -0.6O19879 O.O2273596 O.O35270589 sample clone Sh765B-TZT 20 bacterium Firmicutes Onclassified 3289 Isobacilium meis CCUG. -0.6O2O217 O.O2272631 O.O35270589 3766OT Firmicutes Clostridiaceae 4475 termite gut homogenate -0.6038353 O.O22213S1 O.O35270589 clone RS-NO2 bacterium Firmicutes Enterococcaceae 3598 Enterococcus Sofitarius str. -0.6071271 O.O213OSO6 O.O35270589 DSM S634 Firmicutes Clostridiaceae 4310 termite gut clone RS-056 -0.6O72S21 O.O2127112 O.O35270589 Firmicutes Enterococcaceae 3680 Melissococcus plutonius str. -0.609524 O.O2O66141 O.O35270589 NCDO 2440 Firmicutes Peptococc/ 150 -0.6.166294 O.O1883948 O.O35270589 Acidaminococc Chlorobi Onclassified 549 benzene-degrading nitrate- -0.6183396 O.O1841973 O.O35270589 reducing consortium clone Cart-N2 bacterium Firmicutes Lactobacillaceae 3526 Laciobacilius Sakei -0.6.198.186 O.O1806246 O.O35270589 Firmicutes Lachnospiraceae 4281 granular sludge clone -0.6.271224 O.O1637464 O.O35270589 UASB brew B86 Bacteroidetes Flavobacteriaceae 5726 Bergeyella sp. oral AK152 -0.628.1038 O.O1615736 O.O35270589 clone Firmicutes Lachnospiraceae 4512 granular sludge clone -0.632OO73 O.O1531468 O.O35270589 UASB brew B25 Firmicutes Lachnospiraceae 4331 granular sludge clone -0.6379284 O.O1410088 O.O35270589 UASB brew B84 Firmicutes Lactobacillaceae 3547 Lactobacilius frumenti str. -0.5324,516 O.04998019 O.O35270589 TMW 1.666

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SEQUENCE LISTING

<16O is NUMBER OF SEO ID NOS: 10

<210s, SEQ ID NO 1 &211s LENGTH: 2O &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 1 actic ctacgg gaggcagcag

<210s, SEQ ID NO 2 &211s LENGTH: 17 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 2 attaccgcgg Ctgctgg 17

<210s, SEQ ID NO 3 &211s LENGTH: 22 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 3 gaacggaaag gcc ctgcttg Ca 22

<210s, SEQ ID NO 4 &211s LENGTH: 22 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 4 ggct cotlatc cqg tattaga cc 22

<210s, SEQ ID NO 5 &211s LENGTH: 25 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 5 ggtaaaggct caccalagacic gtgat 25

<210s, SEQ ID NO 6 &211s LENGTH: 2O &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 6 t cacgcggcg ttgctic catc

<210s, SEQ ID NO 7 US 2016/0310546 A1 Oct. 27, 2016

- Continued

&211s LENGTH: 18 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OO > SEQUENCE: 7 tgttggcggga aagacagc 18

<210s, SEQ ID NO 8 &211s LENGTH: 23 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 8 cct tcc tatg gcttagctitc agc 23

<210s, SEQ ID NO 9 &211s LENGTH: 25 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 9 caccacacct tctacaatga gctgc 25

<210s, SEQ ID NO 10 &211s LENGTH: 25 &212s. TYPE: DNA <213> ORGANISM: Artificial Sequence 22 Os. FEATURE: <223> OTHER INFORMATION: Synthetic polynucleotide

<4 OOs, SEQUENCE: 10 acaccctgga tagcaacgta catgc 25

What is claimed is: 7. The method of claim 1, wherein the pharmaceutical 1. A method of treating sinusitis in an individual in need composition is administered to a sinonasal mucosa in the thereof comprising nasally administering a pharmaceutical individual. composition comprising an effective amount of Lactobacil 8. A method of treating sinusitis in an individual in need lus sakei; wherein the sinusitis comprises a Corynebacte thereof comprising administering an effective amount of rium sinus infection; thereby treating the sinusitis in the Lactobacillus sakei to the individual, thereby treating the individual. sinusitis in the individual. 2. The method of claim 1, wherein the Corny bacterium is 9. The method of claim 8, wherein the Lactobacillus sakei Corny bacterium tuberculostearicum, Corny bacterium seg colonizes a sinonasal mucosa of the individual. mentosum, or a combination thereof. 10. The method of claim 9, wherein the sinonasal mucosa 3. The method of claim 1, wherein the Lactobacillus sakei is the maxillary sinus. colonizes the sinonasal mucosa of the individual. 11. The method of claim 8, wherein the sinusitis com 4. The method of claim 3, wherein the sinonasal mucosa prises a Corynebacterium sinus infection. is the maxillary sinus. 12. The method of claim 11, wherein the Corny bacterium 5. The method of claim 1, wherein the pharmaceutical is Corny bacterium tuberculostearicum, Corny bacterium composition further comprises a second bacteria from the segmentosum, or a combination thereof. phylum Firmicutes and family Streptococcaceae. 13. The method of claim 8, comprising nasally adminis 6. The method of claim 5, wherein the second bacteria tering the Lactobacillus sakei to the individual. from the phylum Firmicutes and family Streptococcaceae is 14. The method of claim 8, wherein the pharmaceutical Streptococcus constellatus, Streptococcus mutans, Strepto composition is administered trans-nasally or to the sinuses. coccus suis, Streptococcus thermophiles, Lactococcus lactis, 15. The method of claim 8, wherein the Lactobacillus Streptococcus downei, Streptococcus cristatus, or a combi sakei is administered to the sinonasal mucosa in the indi nation of two or more thereof. vidual. US 2016/0310546 A1 Oct. 27, 2016 22

16. The method of claim 8, further comprising adminis tering a second bacteria from the phylum Firmicutes and family Streptococcaceae. 17. The method of claim 16, wherein the second bacteria from the phylum Firmicutes and family Streptococcaceae is Streptococcus constellatus, Streptococcus mutans, Strepto coccus suis, Streptococcus thermophiles, Lactococcus lactis, Streptococcus downei, Streptococcus cristatus, or a combi nation of two or more thereof. 18. The method of claim 8, further comprising detecting the microbial diversity of the sinus mucosa of the individual. 19. The method of claim 8, further comprising detecting the relative level of Corynebacterium in a mucosal sample from the individual, and comparing the relative level of Corynebacterium in the sample to a control of Corynebac terium levels. 20. The method of claim8, wherein the sinusitis is chronic sinusitis.