Microbiome profiling of uncinate tissue and nasal polyps in patients with chronic rhinosinusitis using swab and tissue biopsy

Chronic rhinosinusitis (CRS) is characterized according to the presence or absence of nasal polyps (NPs) and displays nasal microbiota dysbiosis. However, optimal sampling methods of the nasal microbiome in CRS have not been identified. We aimed to assess the microbial composition in patients with CRS, comparing different sampling methods (swab and tissue biopsy), tissue types (uncinate tissue and NP), and disease subtypes. Samples were obtained by swabbing the middle meatus and taking a biopsy of uncinate tissue (UT) in patients with CRS with (CRSwNP, N = 8) or without NP (CRSsNP, N = 6) and controls (N = 8). NPs were also harvested in CRSwNP. DNAs were extracted from fifty-two samples and analyzed by 16S rRNA gene amplicon sequencing. As a result, a great interpersonal variance was observed in nasal swabs, while UT samples presented distinct microbiome with low inter-personal differences. Moreover, the UT microbiomes were further differentiated into three clusters which are associated with disease status (control, CRSsNP, and CRSwNP). Compared to UT, NP revealed a unique microbiome profile with significantly less bacterial diversity. Prevotella was the genus whose abundance was negatively correlated with disease severity in NP. In conclusion, tissue samples are better specimens than nasal swabs for assessing the microbiomes of CRS patients. Several bacteria in UT and NP tissues revealed an association with clinical severity of CRSwNP.

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Microbiome profiling reveals distinct microbial compositions of nasal polyps in chronic rhinosinusitis

10.21203/rs.3.rs-58912/v1 ◽

2020 ◽

Author(s):

Sung-Woo Cho ◽

Dong-Young Kim ◽

Sungmi Choi ◽

Hye-Ryun Kang ◽

Hana Yi

Keyword(s):

Chronic Rhinosinusitis ◽

Nasal Polyps ◽

Sampling Methods ◽

Clinical Severity ◽

Rrna Gene ◽

Microbial Composition ◽

Principal Coordinates Analysis ◽

Principal Coordinates ◽

Nasal Swabs ◽

Disease Subtypes

Abstract Background: Chronic rhinosinusitis (CRS) is characterized according to the presence or absence of nasal polyps (NPs) and displays nasal microbiota dysbiosis. However, optimal sampling methods of the nasal microbiome in CRS have not been identified. We therefore aimed to assess the microbial composition in patients with CRS, comparing different sampling methods (swab and tissue biopsy), tissue types (uncinate tissue and NP), and disease subtypes.Methods: Samples were obtained by swabbing the middle meatus and taking a biopsy of uncinate tissue (UT) in 8 patients with CRS with NP (CRSwNP), 6 patients with CRS without NP (CRSsNP), and 8 controls. NPs were also harvested in CRSwNP. Extracted DNA samples were analyzed by 16S rRNA gene amplicon sequencing.Results: The microbiome diversity did not significantly differ between disease subtypes in nasal swabs or UT samples. However, a principal coordinates analysis based on weighted UniFrac distances revealed a greater interpersonal variance for nasal swabs than for UT. UT samples presented with a distinct pattern and composition in CRSwNP compared to CRSsNP or control. Compared to UT, NP revealed a unique microbiome profile with significantly less bacterial diversity. Prevotella was the most abundant genus in the UT regardless of the disease subtype and its prevalence was significantly reduced in NP. Prevotella abundance was negatively correlated with disease severity as measured by Lund-Mackay scoring.Conclusion: Tissue samples are better specimens than nasal swabs for assessing the microbiomes of CRS patients. Microbiomes of NP tissues revealed an association with clinical severity of CRSwNP.

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Comparison of Subtyping Approaches and the Underlying Drivers of Microbial Signatures for Chronic Rhinosinusitis

mSphere ◽

10.1128/msphere.00679-18 ◽

2019 ◽

Vol 4 (1) ◽

Cited By ~ 7

Author(s):

Kristi Biswas ◽

Raewyn Cavubati ◽

Shan Gunaratna ◽

Michael Hoggard ◽

Sharon Waldvogel-Thurlow ◽

...

Keyword(s):

Tight Junction ◽

Chronic Rhinosinusitis ◽

Inflammatory Markers ◽

Nasal Polyps ◽

Epithelial Barrier ◽

Clear Understanding ◽

Microbial Composition ◽

Data Set ◽

Heterogeneous Condition ◽

Gene And Protein Expression

ABSTRACT Chronic rhinosinusitis (CRS) is a heterogeneous condition characterized by persistent sinus inflammation and microbial dysbiosis. This study aimed to identify clinically relevant subgroups of CRS patients based on distinct microbial signatures, with a comparison to the commonly used phenotypic subgrouping approach. The underlying drivers of these distinct microbial clusters were also investigated, together with associations with epithelial barrier integrity. Sinus biopsy specimens were collected from CRS patients (n = 23) and disease controls (n = 8). The expression of 42 tight junction genes was evaluated using quantitative PCR together with microbiota analysis and immunohistochemistry for measuring mucosal integrity and inflammation. CRS patients clustered into two distinct microbial subgroups using probabilistic modelling Dirichlet (DC) multinomial mixtures. DC1 exhibited significantly reduced bacterial diversity and increased dispersion and was dominated by Pseudomonas, Haemophilus, and Achromobacter. DC2 had significantly elevated B cells and incidences of nasal polyps and higher numbers of Anaerococcus, Megasphaera, Prevotella, Atopobium, and Propionibacterium. In addition, each DC exhibited distinct tight junction gene and protein expression profiles compared with those of controls. Stratifying CRS patients based on clinical phenotypic subtypes (absence or presence of nasal polyps [CRSsNP or CRSwNP, respectively] or with cystic fibrosis [CRSwCF]) accounted for a larger proportion of the variation in the microbial data set than with DC groupings. However, no significant differences between CRSsNP and CRSwNP cohorts were observed for inflammatory markers, beta-dispersion, and alpha-diversity measures. In conclusion, both approaches used for stratifying CRS patients had benefits and pitfalls, but DC clustering provided greater resolution when studying tight junction impairment. Future studies in CRS should give careful consideration to the patient subtyping approach used. IMPORTANCE Chronic rhinosinusitis (CRS) is a major human health problem that significantly reduces quality of life. While various microbes have been implicated, there is no clear understanding of the role they play in CRS pathogenesis. Another equally important observation made for CRS patients is that the epithelial barrier in the sinonasal cavity is defective. Finding a robust approach to subtype CRS patients would be the first step toward unravelling the pathogenesis of this heterogeneous condition. Previous work has explored stratification based on the clinical presentation of the disease (with or without polyps), inflammatory markers, pathology, or microbial composition. Comparisons between the different stratification approaches used in these studies have not been possible due to the different cohorts, analytical methods, or sample sites used. In this study, two approaches for subtyping CRS patients were compared, and the underlying drivers of the heterogeneity in CRS were also explored.

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PCR Analysis of Nasal Polyps, Chronic Sinusitis, and Hypertrophied Turbinates for DNA Encoding Bacterial 16S rRNA

American Journal of Rhinology ◽

10.1177/194589240201600309 ◽

2002 ◽

Vol 16 (3) ◽

pp. 169-173 ◽

Cited By ~ 21

Author(s):

Gerald A Bucholtz ◽

Sherry A. Salzman ◽

Fernando B. Bersalona ◽

Timothy R. Boyle ◽

Victor S. Ejercito ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Nasal Polyps ◽

Chronic Sinusitis ◽

Consensus Sequence ◽

Rrna Gene ◽

Dna Encoding ◽

Bacterial Dna ◽

Two Samples ◽

The 16S Rrna Gene

Background Nasal polyps are considered to result from chronic inflammation, but the initial or persisting stimulus for the inflammation is not known. A variety of bacteria and fungi have been cultured from nasal polyps, but ∼35% have sterile cultures. Previously, Mycoplasma pneumoniae–specific DNA was detected in human nasal polyps using polymerase chain reaction (PCR) techniques, suggesting M. pneumoniae as a causative agent in the etiology of nasal polyps. Methods In this study, we tested for the presence of bacterial DNA in nasal polyps resected from 40 patients, in nasal mucosa membrane from 9 patients undergoing turbinectomy for hypertrophy, and in sinus mucosa membrane from 6 patients undergoing endoscopic surgery for chronic sinusitis. Tissue DNA was extracted and analyzed by PCR using M. pneumoniae specific primers for DNA that encode the 16S rRNA gene in 41 specimens (31 polyps, 6 turbinates, and 4 sinus), and by consensus sequence-based PCR using broad range primers for most eubacterial DNA encoding the 16S rRNA gene in 38 specimens (26 polyps, 7 turbinates, and 5 sinuses). Results Only two samples were positive for bacterial DNA encoding 16S rRNA: Streptococcus sp. DNA was isolated from one polyp specimen and Pseudomonas aeruginosa DNA was isolated in one maxillary sinusitis specimen. No evidence of M. pneumoniae–specific DNA encoding 16S rRNA was found in any of the tissues. Conclusions This study suggests that chronic bacterial infection is not a major component of nasal polyp etiology.

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2356 The nasopharyngeal microbiome is perturbed and associated with increased clinical severity during acute respiratory viral infection

Journal of Clinical and Translational Science ◽

10.1017/cts.2018.137 ◽

2018 ◽

Vol 2 (S1) ◽

pp. 32-32

Author(s):

Darrell Dinwiddie ◽

Ashlee K. Bradley ◽

Jesse L. Denson ◽

Joshua L. Kennedy ◽

Walter N. Dehority ◽

...

Keyword(s):

Viral Infections ◽

Severe Disease ◽

Clinical Severity ◽

Therapeutic Interventions ◽

Rrna Gene ◽

Microbial Composition ◽

Mixed Cell ◽

Severe Acute Respiratory Infection ◽

Syncytial Virus

OBJECTIVES/SPECIFIC AIMS: We sought to investigate the role of the host microbiome during severe, acute respiratory infection (ARI) to understand the drivers of both acute clinical pathogenesis. METHODS/STUDY POPULATION: Nasopharyngeal swabs comprised of mixed cell populations at the active site of infection were collected from 192 hospitalized pediatric patients with ARI. We combined comprehensive respiratory virus detection and virus genome sequencing with 16S rRNA gene sequencing to evaluate the microbial content of the airway during ARI. This data was coupled with 11 clinical parameters, which were compiled to create a clinical severity score. The microbiome profiles were assessed to determine if clinical severity of infection, and/or specific virus was associated with increased clinical severity. RESULTS/ANTICIPATED RESULTS: We identified 8 major microbiome profiles classified by dominant bacterial genus, Moraxella, Corynebacterium, Staphylococcus, Haemophilus, Streptococcus, Alloiococcus, Schlegelella, and Diverse. Increased clinical severity was significantly associated with microbiome profiles dominated by Haemophilus, Streptococcus, and Schlegelella, whereas Corynebacterium and Alloiococcus were more prevalent in children with less severe disease. Independent of the microbial community, more than 60% of patients with the highest clinical severity were infected with either respiratory syncytial virus or rhinovirus. DISCUSSION/SIGNIFICANCE OF IMPACT: Our results indicate that individually and in combination, both virus and microbial composition may drive clinical severity during acute respiratory viral infections. It is still unclear how the complex interplay between virus, bacterial community, and the host response influence long-term respiratory impacts, such as the development of asthma. Nonetheless, during ARIs therapeutic interventions such as antibiotics and probiotics may be warranted in a subset of patients that are identified to have both a virus and microbiome profile that is associated with increased pathogenesis to limit both acute and long-term phenotypes.

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Heat Shock Protein 70 and Anti-heat Shock Protein 70 Antibodies in Nasal Secretions of Patients with Chronic Rhinosinusitis

Allergy & Rhinology ◽

10.2500/ar.2016.7.0149 ◽

2016 ◽

Vol 7 (1) ◽

pp. ar.2016.7.0149 ◽

Cited By ~ 2

Author(s):

Namjil N. Tsybikov ◽

Elena V. Egorova ◽

Boris I. Kuznik ◽

Elena V. Fefelova ◽

Eli Magen

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Chronic Rhinosinusitis ◽

Heat Shock Protein 70 ◽

Nasal Polyps ◽

Immunoglobulin E ◽

Clinical Severity ◽

Secretory Iga ◽

Enzyme Linked Immunosorbent Assay ◽

Antibody Levels

Background The issue of heat shock protein (HSP) 70 and anti-HSP70 antibodies in chronic rhinosinusitis (CRS) has never been explored. Objective To determine the nasal secretion (NS) levels of HSP70 and anti-HSP70 antibodies in patients with CRS with nasal polyps (CRSwNP) and patients with CRS without nasal polyps (CRSsNP), and to evaluate their associations with CRS clinical severity and correlation with NS interleukin (IL), IL-5 and interferon λ. Methods CRS severity was determined by Lund-Mackay scores. Levels of immunoglobulin E (IgE), IL-4, IL-5, interferon A, HSP70, and anti-HSP70 antibody levels in NS were measured by enzyme-linked immunosorbent assay. Results Forty-six patients with CRSsNP (25 women [543%] and 21 men [45.7%], mean [standard deviation {SD}]) age, 34.1 ± 123 years; 54 patients with CRSwNP (24 women [44.4%] and 30 men [55.6%], mean [SD] age, 37.9 ± 17.5 years). A group of 40 healthy subjects served as controls. Compared with the controls (with a mean [SD] NS HSP70 level of 0.05 ± 0.03 μg/mL), mean [SD]NS HSP70 levels in both the CRSsNP group (0.16 ± 0.07 ixg/mL) and CRSwNP group (0.21 ± 0.10 μg/mL) were increased (p < 0.001). Similarly, the mean (SD) NS anti-HSP70 antibody levels were significantly higher in patients with CRSwNP (0.25 ± 0.09 optical density value [ODV]) compared with CRSsNP (0.13 ± 0.04 ODV) (p < 0.001) and healthy controls (0.14 ± 0.02 ODV) (p < 0.001). NS HSP70 in subjects with CRSwNP showed a significant positive correlation with the Lund-Mackay score (r = 0.31; p < 0.05). NS levels of either HSP70 or anti-HSP70 antibodies were strongly correlated with NS IL-4 in the CRSwNP group (r = 0.62, p < 0.001; and r = 0.69, p < 0.001, respectively). Conclusion NS concentrations of HSP70 and secretory IgA anti HSP70 antibodies are increased in CRSwNP (but not in CRSsNP) and correlate positively with the Lund-Mackay score, NS IL-4, and NS IL-5.

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IL-33 Expression in Chronic Rhinosinusitis with Nasal Polyps and Its Relationship with Clinical Severity

ORL ◽

10.1159/000484527 ◽

2017 ◽

Vol 79 (6) ◽

pp. 323-330 ◽

Cited By ~ 7

Author(s):

Wei Song ◽

Chunlei Wang ◽

Jing Zhou ◽

Shenghua Pan ◽

Sen Lin

Keyword(s):

Chronic Rhinosinusitis ◽

Nasal Polyps ◽

Clinical Severity

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Mycoplasma pneumoniaeandChlamydophila pneumoniae: a comparative study in patients with nasal polyposis and healthy controls

The Journal of Laryngology & Otology ◽

10.1017/s0022215115001590 ◽

2015 ◽

Vol 129 (9) ◽

pp. 865-869 ◽

Cited By ~ 1

Author(s):

D G Ioannidis ◽

V A Lachanas ◽

Z Florou ◽

J G Bizakis ◽

E Petinaki ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Real Time ◽

Chronic Rhinosinusitis ◽

Nasal Polyps ◽

Inferior Turbinate ◽

Sinus Surgery ◽

Control Group ◽

Chain Reaction ◽

Tissue Samples ◽

Polymerase Chain

AbstractIntroduction:The role played byMycoplasma pneumoniaeandChlamydophila pneumoniaein the pathogenesis of chronic rhinosinusitis with nasal polyps has been the object of ongoing debate. We used real-time polymerase chain reaction to investigate the prevalence of both microorganisms in the nasal tissue samples of patients and controls.Methods:We extracted DNA from nasal polyp samples obtained during functional endoscopic sinus surgery and the inferior turbinate samples of controls undergoing septoplasty. We used the highly sensitive real-time polymerase chain reaction to detect the presence ofM pneumoniaeandC pneumoniaeDNA.Results:Patients with chronic rhinosinusitis with nasal polyps consisted of 62 individuals (39 men; mean age 51 years); the control group consisted of 24 individuals (13 men; mean age 45 years). All samples from both groups were negative forM pneumoniaeandC pneumoniaeDNA.Conclusion:We have demonstrated that the likelihood ofM pneumoniaeandC pneumoniaeacting as an ongoing inflammatory stimulus in chronic rhinosinusitis with nasal polyps is slim.

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No evidence for a correlation of glutathione S-tranferase polymorphisms and chronic rhinosinusitis

Rhinology Journal ◽

10.4193/rhino10.064 ◽

2011 ◽

Vol 49 (2) ◽

pp. 180-184

Author(s):

K. Fruth ◽

N. Best ◽

M. Amro ◽

K. Ingel ◽

J. Gosepath ◽

...

Keyword(s):

Oxidative Stress ◽

Chronic Rhinosinusitis ◽

Nasal Polyps ◽

Bronchial Hyperresponsiveness ◽

Inferior Turbinate ◽

Tissue Samples ◽

Cellular Protection ◽

Functional Variants ◽

Cellular Detoxification ◽

The Individual

OBJECTIVE: Cellular detoxification mechanisms are mandatory for cellular protection against oxidative stress and reactive oxygen species. One major group of antioxidative active enzymes involved in cellular detoxification are the Glutathione S-Transferases (GST). Multiple subtypes like GSTM1, GSTP1, and GSTT1 and variants of them are known, arising from allelic variations of the GST loci. Moreover, functional variants occur in high percentages and have been associated with diseases like bronchial asthma and bronchial hyperresponsiveness. The interplay of oxidative stress, detoxifying genes like GSTs and the genesis of respiratory tract illness is under contradictory debate. In this study, we analysed the potential association of GST-polymorphisms and chronic rhinosinusitis (CRS). METHODS: In total 170 nasal tissue samples, 49 tissue samples from patients with CRS without nasal polyps, 69 tissue samples from CRS with nasal polyps and 52 healthy tissue controls of the inferior turbinate were analysed for their individual GST-status. Genotypes for GSTM1 (null versus present), GSTT1 (null versus present), and GSTP1 (Ile105Val) were determined by Polymerase Chain Reaction. The respective genotypes were correlated to the incidence of CRS with and without nasal polyps in aspirin-tolerant and intolerant patients and to the individual health status concerning asthma and allergies. RESULTS: No correlation between any GST-polymorphism and CRS with and without nasal polyps or allergies or asthma or aspirin-intolerance was observed. CONCLUSION: Our results do not suggest that there is a relevant genetic predisposition considering the individual GST-status for the susceptibility of nasal respiratory epithelia leading to CRS.

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IL-19 Induced by IL-13/IL-17A in the Nasal Epithelium of Patients with Chronic Rhinosinusitis Upregulates MMP-9 Expression via ERK/NF-kB Signaling Pathway

10.21203/rs.3.rs-46150/v3 ◽

2020 ◽

Author(s):

Xia Li ◽

Jiancong Huang ◽

Xiaohong Chen ◽

Xiaoping Lai ◽

Zizhen Huang ◽

...

Keyword(s):

Western Blot ◽

Chronic Rhinosinusitis ◽

Nasal Polyps ◽

Tissue Remodeling ◽

Tissue Inhibitors Of Metalloproteinases ◽

Tissue Samples ◽

Nasal Tissue ◽

Human Nasal Epithelial Cells ◽

Pathway Activation

Abstract Background: Tissue remodeling is a crucial characteristic of chronic rhinosinusitis (CRS). Imbalance between matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) is crucial for the pathologic tissue remodeling in CRS. Elevation of interleukin (IL)-19 or MMP-9 levels in patients with CRS had been proven in previous studies. Here, we aimed to investigate the role of IL-19 in mediating MMP-9 expression in CRS. Methods: Nasal tissue samples were collected from 45 individuals having chronic rhinosinusitis with nasal polyps (CRSwNP), 24 CRS without nasal polyps (CRSsNP), and 17 controls. Expression of IL-19, its receptors (IL-20R1/IL-20R2), and MMP-9 were investigated using RT-qPCR and Immunofluorescence. Human nasal epithelial cells (HNECs) were stimulated by IL-19; ERK phosphorylation, NF-kB pathway activation, and MMP-9 level were detected by RT-qPCR, ELISA, western blot and Immunofluorescence. We also explored the effect of type1/2/3 cytokines on IL-19 production by RT-qPCR, and western blot. Results: Expression levels of IL-19, its receptors (IL-20R1/IL-20R2), and MMP-9 were increased in nasal tissues from individuals with CRSwNP compared to those with CRSsNP as well as the controls. IL-19 significantly elevated the production of MMP-9 in HNECs. Furthermore, IL-19 could activate the ERK and NF-kB pathways, accompanied by increased MMP-9 production in HNECs. Conversely, both ERK and NF-kB inhibitors significantly attenuated the role of IL-19 in MMP-9 production. siRNA knockdown of IL-20R1 suppressed ERK and NF-kB pathway activation, thereby decreasing MMP-9 expression. IL-13 and IL-17A were found to stimulate IL-19 production in HNECs.Conclusion: IL-19, promoted by IL-13 and IL-17A, contributes to the upregulation of secretion of the tissue remodeling factor MMP-9 in patients with CRS.

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