CN110218784B - Application of flora abundance detector in preparation of nasal polyp and prognosis detection agent thereof - Google Patents
Application of flora abundance detector in preparation of nasal polyp and prognosis detection agent thereof Download PDFInfo
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Abstract
The invention relates to the field of nasal polyps and prognosis detection thereof, in particular to application of a flora abundance detector in preparation of nasal polyps and a prognosis detection agent thereof. The present invention is directed to assessing the relationship between sinus microorganism composition and inflammation type by comparing the differences in sinus microorganism composition and structure between CRSwNP and healthy controls; to determine if certain sinus microbial compositions are associated with post-operative polyp recurrence in CRSwNP patients, the synergistic predictive value of inflammatory cells and differential bacteria for polyp recurrence is further evaluated.
Description
Technical Field
The invention relates to the field of nasal polyps and prognosis detection thereof, in particular to application of a flora abundance detector in preparation of nasal polyp typing and a prognosis detection agent thereof.
Background
Chronic rhinosinusitis with nasal polyps (CRSwNP) is a complex and heterogeneous inflammation of the mucous membranes of the sinuses characterized by the presence of nasal polyps estimated to occur in 1% to 4% of the world population. In europeans and americans, CRSwNP is characterized immunologically predominantly by Th2 inflammation; whereas in asian countries such as china and korea, at least half of CRSwNP patients exhibit Th1/Th17 inflammatory responses. Inflammation typing is based on the proportion of inflammatory cells that infiltrate the tissue, eosinophilia being one of the most critical features. Th2 inflammation is associated with recurrence of nasal polyps. Eosinophils are the most effective markers for predicting recurrence of nasal polyps.
That is, previous studies have considered that pre-operative CT score, nasal polyp score, eosinophil, combined asthma or allergic status, interleukin-5 expression, adhesion molecule expression could predict recurrence of post-operative nasal polyp, where tissue eosinophil percentage is the most effective marker of these indicators. However, these indicators have certain limitations in predicting recurrence of nasal polyps. Most indexes are detected in nasal polyp tissue samples, and the acquisition of nasal polyps is an invasive method, so that not only can the nasal bleeding be increased, the fear of a patient is caused, but also the risk of infection is increased. More importantly, these indices through nasal polyp sample detection are often used to predict efficacy after intranasal endoscopic surgery, and are of little significance in the selection of pre-operative drug treatments and surgical protocols.
In view of this, the present invention has been made.
Disclosure of Invention
The aim of the present study was to 1) compare the differences in sinus microorganism composition and structure between CRSwNP and healthy controls, in particular to evaluate the relationship between sinus microorganism composition and inflammation type; 2) To determine if certain sinus microbial compositions are associated with post-operative polyp recurrence in CRSwNP patients, the synergistic predictive value of inflammatory cells and differential bacteria for polyp recurrence is further evaluated.
In order to achieve the above object of the present invention, the following technical solutions are specifically adopted:
in one aspect, the invention provides an application of a flora abundance detector in preparing a nasal polyp detection agent.
Further, the flora comprises phylum level bacteria and/or genus level bacteria;
the portal level bacteria are any one or more of the following: proteobacteria, actinobacteria, bacteroidetes, firmicutes, fusobacteria;
the genus level bacteria are any one or more of the following: corynebacterium Staphylococcus, veillonella, escherichia/Shigella, streptococcus.
It has been found that in addition to the immunoinflammatory changes of tissue, paranasal sinus microorganisms, pathogens and their metabolites may also play a role in the pathogenesis of CRSwNP. Previous studies have observed that possible mechanisms by which sinus microorganisms promote nasal polyp occurrence include reduced microbiota diversity; with pathogen exposure, disruption of the epithelial barrier may further promote inflammatory responses by specific bacterial colonization or bacterial biofilm formation. However, few studies investigate the sinus microbiome profile in CRSwNP patients with different inflammation types, as well as the association with clinical features and/or polyp recurrence.
The invention extracts the microbial genome from the nasal passage falling object obtained by the nasal passage cotton swab of the patients with different types of chronic nasosinusitis and nasal polyp and the normal people, detects the V3-V4 region of the bacterial 16S ribosomal RNA gene, and sequences the product to analyze the abundance of the thalli. Specifically, OTU clustering was performed at 97% similarity using UPARSE, and chimeric sequences were identified and removed using Userach (version 7.0). Each representative sequence was taxonomically identified by a Ribosome Database Project (RDP) classifier in the RDP database (http:// RDP. Cme. Msu. Edu /), using a confidence threshold of 0.8. OTU analysis Table and alpha/beta diversity analysis were performed by QIIME.
It was found that CRSwNP flora was reduced in abundance and uniformity (α diversity) compared to the normal control group, and that the inter-sample flora structure was greatly altered (β diversity) compared to the normal control group. Further typing studies, especially neutrophil nasal polyps, showed a marked decrease in bacterial alpha diversity and a maximal variation in flora structure (beta diversity) compared to the normal control group.
In terms of flora abundance, the common flora is at the portal level Proteobacteria, bacterioides, firmics, fusobacteria, actinomycetes, with bacterial abundance exceeding 90%. Compared to the control group, CRSwNP increased in Proteobacteria abundance, while bacteria, firmicutes and Fusobacteria decreased in abundance. In contrast, in the different typing studies, neutrophil nasal polyps were significantly increased compared to eosinophil nasal polyps and controls. At the genus level, most of the probiotics of CRSwNP are decreasing in abundance and pathogenic bacteria are increasing in abundance. In typing studies, neutrophil nasal polyps have more corynebacteria and Staphylococcus, less Escherichia/Shigella, veillonella, streptococcus.
Relationship between flora and clinical index: tissue neutrophil% and Acinetobacter actylobacter, belonging to the genus Leptobacter, being positively correlated with Staphylococcus, and being negatively correlated with Bacillus, prevotella, escherichia/Shigella, streptococcus and Veillonella. The smell is inversely related to the Acinetobacter of the door level bacteria, belonging to the genus Corynebacterium, staphylococcus and Veilonella. Runny nose is positively correlated with a genus level of bacteria Pseudomonas.
In another aspect, the invention provides the use of a flora abundance detector in the preparation of a nasal polyp prognosis detector.
The indexes studied in the past have a certain limitation in predicting the recurrence of nasal polyps, and most indexes are detected in nasal polyp tissue samples, and the acquisition of the nasal polyps is an invasive method, so that not only can the nasal bleeding be increased, but also the fear of patients can be increased, and the risk of infection can be increased. More importantly, these indices through nasal polyp sample detection are often used to predict efficacy after intranasal endoscopic surgery, and are of little significance in the selection of pre-operative drug treatments and surgical protocols.
Although Actinobacteria (Corynebacterium) is considered to be related to the efficacy of the nasal sinusitis after surgery by the existing research, nasal bacteria Actinobacteria (Corynebacterium) are considered to be high in abundance before sinusitis surgery, which is good in efficacy; there are also studies demonstrating that Actinobacteria (Corynebacterium) is less abundant in CRS with allergic rhinitis than CRS with less allergic rhinitis. While the abundance of bacterium Actinobacteria (Corynebacterium) is inversely related to the cytokine IL-5, ECP, has also been found by recent studies. However, no correlation between flora and nasal polyp typing and prognosis has been reported in the prior art.
Further, the flora comprises phylum level bacteria and/or genus level bacteria;
the portal level bacteria are any one or more of the following: proteobacteria, actinobacteria, bacteroidetes, firmicutes, fusobacteria;
the genus level bacteria are any one or more of the following: staphylococcus, corynebacterium, prevotella, fusobacterium, clostridium sensu stricto, veillonella, corynebacterium, staphylococcus, propionibacterium, anaerococcus.
Preferably, the portal level bacteria are actinomycetes and/or Proteobacteria.
Preferably, the genus-level bacterium is Corynebacterium.
The invention obtains the factor capable of predicting nasal polyp recurrence through comparing clinical indexes and flora abundance of recurrent nasal polyp and single-factor and multi-factor binary Logistic regression analysis.
The present study suggests that bacterial Actinobacteria (Corynebacterium) abundance is positively correlated with mucosal neutrophil% and significantly increased in neutrophil nasal polyps. Actinobacteria (Corynebacterium) affects the mechanism of tissue inflammation, probably by promoting benign colonization of staphylococcus aureus and rendering it non-pathogenic, and promotes the conversion of inflammation to Th1/Th17 responses.
In the application of the invention, the detection template of the flora abundance is a microbial genome extracted from the middle nasal meatus drop.
Further, a middle meatus cotton swab is used to obtain a middle meatus falling object.
Further, the nasal polyp is chronic rhinosinusitis with nasal polyp.
In the above application of the present invention, the detection agent further comprises a reagent for detecting the percentage of eosinophils in the tissue.
In the above application of the present invention, the detection object includes any one or more of a primer, a probe, a biosensor and a chip, but is not limited thereto, and the technical means for detecting the diversity of the bacterial flora is within the scope of the present invention.
Further, the test object is designed for the V3-V4 region of the bacterial 16S ribosomal RNA gene.
For example, the primer is shown as SEQ ID NO. 1-2.
The invention also provides a kit for detecting nasal polyp, which comprises a detection object and a reagent for detecting the percentage of eosinophils in tissues;
the test is designed for the V3-V4 region of the bacterial 16S ribosomal RNA gene or for any one or more of the Actinobacteria, corynebacterium flora.
Further, the detection object includes any one or more of a primer, a probe, a biosensor and a chip, but is not limited thereto, and the technical means for detecting the diversity of the bacterial flora is within the scope of the present invention.
The primer is shown as SEQ ID NO. 1-2.
The invention also provides a kit for detecting recurrence of nasal polyp prognosis, which comprises a detection object and a reagent for detecting the percentage of eosinophils in tissues;
the test is designed for the V3-V4 region of the bacterial 16S ribosomal RNA gene or for any one or more of the Actinobacteria, corynebacterium flora.
Further, the detection object includes any one or more of a primer, a probe, a biosensor and a chip, but is not limited thereto, and the technical means for detecting the diversity of the bacterial flora is within the scope of the present invention.
The primer is shown as SEQ ID NO. 1-2.
Compared with the prior art, the invention has the beneficial effects that:
(1) The invention provides the flora abundance for the first time for detecting nasal polyp and detecting the prognosis situation thereof.
(2) The invention obtains specific flora abundance through a large amount of test data analysis, and has a great effect on predicting whether nasal polyps recur or not.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below.
FIG. 1 is a Wen diagram showing the composition of chronic sinusitis nasal polyps (CRSwNP) and health Control (Control) group (A) OTU according to example 1 of the present invention; (B) Shannon index comparison plot;
FIG. 2 is a Wen diagram of three immunophenotyping and health Control (Control) groups (A) OTU compositions of chronic sinusitis with nasal polyps (CRSwNP) according to example 1 of the present invention; (B) Shannon index comparison plot;
FIG. 3 is a graph showing the correlation of the percentage of neutrophils in polyp tissue with Shannon index in example 1 of the present invention;
FIG. 4 is a graph showing principal coordinate analysis of three immunophenotyping and health Control (Control) groups of chronic sinusitis with nasal polyps (CRSwNP) according to example 1 of the present invention;
FIG. 5 is a graph showing the difference between the first principal coordinates (PC 1) and the second principal coordinates (PC 2) of three immunophenotyping and healthy Control (Control) groups of chronic sinusitis and nasal polyps (CRSwNP) according to example 1 of the present invention;
FIG. 6 shows that the percent neutrophils in polyp tissue in example 1 of the present invention are inversely related to the first principal coordinate (PC 1);
FIG. 7 is a bar graph of the abundance of the first 20 high abundance gate-level bacteria of example 1 of the invention;
FIG. 8 is a bar graph of the abundance of the first 20 high abundance genus horizontal bacteria of example 1 of the present invention;
FIG. 9 is a graph showing the difference in abundance of the first 5 high abundance portal level bacteria in chronic rhinosinusitis with nasal polyps (CRSwNP) versus healthy Control (Control) in example 1 of the present invention;
FIG. 10 is a graph showing the difference in abundance between neutrophil-type, eosinophil-type, non-eosinophil/non-neutrophil-type polyps and healthy controls for the first 5 high abundance portal-level bacteria of example 1 of the present invention;
FIG. 11 is a graph showing the difference in bacterial abundance at the level of neutrophil type, eosinophil type, non-eosinophil/non-neutrophil type polyps compared to healthy controls in example 1 of the present invention;
FIG. 12 is a graph showing the correlation of neutrophil percentage and bacterial abundance in polyp tissue in example 1 of the present invention;
FIG. 13 is a graph showing the correlation of smell and bacteria in example 1 of the present invention; correlation of runny nose with bacteria;
FIG. 14 is a bar graph comparing Shannon index for recurrent and non-recurrent groups in chronic sinusitis with nasal polyps (CRSwNP) according to example 2 of the present invention;
FIG. 15 is a principal coordinate analysis of recurrent and non-recurrent groups of nasal polyps with sinusitis (CRSwNP) according to example 2 of the present invention;
FIG. 16 is a bar graph comparing the recurrent group of chronic sinusitis with nasal polyps (CRSwNP) and the non-recurrent group at a first principal coordinate (PC 1) and a second principal coordinate (PC 2) in example 2 of the present invention;
FIG. 17 is a graph showing comparison of the abundance differences between the recurrent and non-recurrent groups of chronic sinusitis and nasal polyps (CRSwNP) at the portal level in example 2 of the present invention;
FIG. 18 is a graph showing comparison of the difference in abundance of bacterial flora at the genus level in a recurrent group and non-recurrent group of chronic sinusitis and nasal polyp (CRSwNP) according to example 2 of the present invention;
FIG. 19 is a graph of the working characteristics (ROC) of a subject in which the door horizontal bacteria Actinobacillus and tissue eosinophil% independently and in combination predict polyp recurrence in example 2 of the present invention;
FIG. 20 is a ROC curve of the horizontal bacteria Corynebacterium and tissue eosinophil% independent and combined prediction of polyp recurrence in example 2 of the present invention.
Detailed Description
Embodiments of the present invention will be described in detail below with reference to examples, but it will be understood by those skilled in the art that the following examples are only for illustrating the present invention and should not be construed as limiting the scope of the present invention. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
Example 1
1. Study object
The study collected nasal cotton swabs (Copan, breccia, italy) from 22 controls and 66 CRSwNP (including 25 eosinophil-type nasal polyps, 14 neutrophil-type nasal polyps, 13 non-eosinophilic/non-neutrophil-type nasal polyps, 14 mixed-type nasal polyps) at beijing co-morat. The control group was derived from patients with anatomic variation for which correction of nasal septum deflection was performed, but without any other nasal disease.
Clinical data collection was performed on all subjects, including sex, age, combined disease (asthma, allergic rhinitis, specificity), preoperative symptom scores (olfactory, nasal obstruction, nasal discharge, headache, sneeze and preoperative total score), baseline level histopathological cytometer percentages (eosinophils, neutrophils, lymphocytes, plasma cells) and preoperative extracellular Zhou Xieyan cell percentages (eosinophils, neutrophils, lymphocytes).
2. Sample collection
Middle nasal tract sampling was performed on CRSwNP patients and control groups under intranasal endoscopic guidance. The sterile cotton swab was gently pressed against the middle nasal meatus mucosa for 30s and the cotton swab was rotated 5 turns under the intranasal scope, then the swab was placed into a sterile collection tube containing transport medium and immediately broken at the red line. Information was labeled and swabs were stored at-80 ℃ until DNA extraction.
3.16S rDNA sequencing
Microbial DNA was extracted from sample cotton swabs using QIAamp DNA Mini Kit (QIAGEN, hilden, germany) according to the protocol of the kit manufacturer. The purity of the DNA was assessed based on A260/A280 using a Thermo NannoDrop 2000 spectrophotometer (Thermo Fisher Scientific, MC, USA). The integrity of the DNA was checked by electrophoresis on a 1% agarose gel.
The V3-V4 region of the bacterial 16S ribosomal RNA gene was amplified by PCR using specific primers (5 '-ACTCCTACGGGRSGCAGCAG) -3' (Forward) and 5 '-GGACTACVVGGGTATCTAATC-3') (95℃for 3 minutes, then 30 cycles, 98℃for 20 seconds, 58℃for 15 seconds, 72℃for 20 seconds and finally primer extension at 72℃for 5 minutes). Sequencing was performed on a HiSeq platform (Illumina, inc., CA, USA) of Realbio GenomicsInstitute (Shanghai, china.) OTU clustering was performed using UPARSE at 97% similarity, and chimeric sequences were identified and removed using Userach (version 7.0.) each representative sequence was identified and classified by a ribosomal database entry (RDP) classifier in the RDP database (http:// rdp.cme.msu.edu /), using a confidence threshold of 0.8. OTU analysis table and alpha/beta diversity analysis were performed by QIIME.
4. Bacterial colony study of different types of chronic nasosinusitis and nasal polyp
Since the mixed form is not typically a single inflammatory CRSwNP, this typing study did not involve mixed nasal polyps.
(1) 2,673,836 sequences and 3310 OTUs were identified.
(2) CRSwNP flora enrichment and uniformity (α diversity) was reduced compared to the normal control group (fig. 1).
Three immunophenotyping of chronic sinusitis with nasal polyps (CRSwNP) showed a decrease in the Shannon index of neutrophilic polyps (Neu) alone compared to the healthy Control (Control) group, indicating a decrease in diversity, whereas eosinophilic polyps (Eos) and non-eosinophilic/non-neutrophilic polyps (Ne/nN) were not significantly different compared to the healthy Control (Control) group (fig. 2). Further typing studies, the percentage of neutrophils in polyp tissue was inversely related to Shannon index, further demonstrating the significantly reduced bacterial diversity of neutrophil nasal polyps (fig. 3).
As shown in fig. 4, there was a difference in principal coordinate analysis between neutrophil type, eosinophil type, and non-eosinophil/non-neutrophil type polyps compared to healthy controls, indicating that the four microorganisms were different in composition. Compared with the normal control group, the neutrophil nasal polyp flora structure variation is the largest (beta diversity).
As shown in fig. 5, neutrophil type, eosinophil type, non-eosinophil/non-neutrophil type polyps exhibited a significant difference in the first and second principal coordinate values compared to the healthy control. As shown in fig. 6, the neutrophil percentage in polyp tissue appears to be inversely related to the first principal coordinate (PC 1).
(3) In terms of flora abundance, fig. 7 shows the abundance of the first 20 high abundance gate-level bacteria; figure 8 shows the first 20 high abundance genus level bacterial abundance cases. At the gate level, the common flora is Proteobacteria, bacterioides, firmics, fusobacteria, actionobacter, with a bacterial abundance ratio of more than 90%. Compared to the control group, CRSwNP increased in Proteobacteria abundance, while bacteria, firmics and Fusobacteria decreased in abundance (fig. 9). Whereas in the different typing studies neutrophil nasal polyps had a significant increase in actionobacter compared to eosinophilic nasal polyps and controls (figure 10).
At the genus level, most of the probiotics of CRSwNP are decreasing in abundance and pathogenic bacteria are increasing in abundance. In typing studies, neutrophil nasal polyps were found to be more Corynebacterium and Staphylococcus, less Escherichia/Shigella, veillonella, streptococcus (fig. 11).
(4) Relationship between flora and clinical index: tissue neutrophil% and Acinetobacter actinomyces, belonging to the genus Horizoctonia Corynebacterium, staphylococcus are positively correlated and to the genus Horizoctoides, belonging to the genus Horizoctodes, prevolella, escherichia/Shigella, streptococcus and Veillonella are negatively correlated (FIG. 12).
The smell was inversely related to the genus Lactobacillus, corynebacterium, staphylococcus, and positively related to the genus Lactobacillus, veilonella, and the nasal discharge was positively related to the genus Lactobacillus, pseudomonas (FIG. 13).
Example 2
Relationship between flora and recurrence of polyp
The study on recurrence included all nasal polyp samples
1. Recurrent and non-recurrent nasal polyps were not significantly different in flora richness and uniformity, but were significantly different throughout the flora structure (fig. 14-16).
2. Recurrence and non-recurrence were different at portal and genus level flora abundance (fig. 17-18). The recurrent group had more of the phylum Proteobacteria, belonging to the genus Leptobacter, fusobacterium, clostridium sensu stricto and Veilonella, less of the phylum Leptobacter Actinobacillus, belonging to the genus Leptobacter, staphylococcus, propionibacterium and Anaerococcus.
3. Flora can predict recurrence of nasal polyp
Incorporating the clinical indicators of the differences and the differential flora into Logistic regression analysis we found that only tissue eosinophils% (the area under the curve (AUC) =0.908) and door horizontal bacteria actionobacteria (auc=0.786) (fig. 19), belonging to horizontal bacteria Corynebacterium (auc=0.811) (fig. 20), could predict recurrence of nasal polyps. Furthermore, the combined organization of eosinophil% and the phylum level bacterium actionobacteria or the genus level bacterium Corynebacterium can significantly increase the predictive value of nasal polyp recurrence (auc=0.962, auc=0.981).
From the above, it is clear that CRSwNP is present with a disturbed flora, which is probably one of the causative mechanisms of CRSwNP pathogenesis. And different flora structures exist between the inflammation types, especially neutrophil type nasal polyps, and the flora colonization and the inflammation types are obviously related. Different flora abundances may be associated with nasal polyp recurrence and may be predictive of polyp recurrence, and combining inflammatory cells may significantly increase the ability to predict polyp recurrence. This study may be instructive for nasal polyp typing and may predict post-operative outcome and adjust medication via flora.
Example 3
Method for predicting recurrence by screening flora
Except for 1 person losing visit, the other 65 persons are subjected to standard follow-up medication, and complete clinical data is provided for 66 nasal polyp patients. Of these, 31 in the recurrent group and 34 in the non-recurrent group.
1. Single factor analysis
The preoperative 17 clinical indexes of the recurrent group and the non-recurrent group are compared, and the relative abundance of the genus level bacteria of the front 5 and the front 30 of the abundance obtained by 16s rDNA sequencing is treated as the statistical difference with the statistical P value of less than 0.05.
The 17 clinical indexes are respectively: age, sex, combined asthma, combined allergic rhinitis, combined atopy, preoperative symptom score (olfactory, nasal obstruction, runny nose, headache, sneeze, total preoperative symptoms), percentage of histoinflammatory cells (eosinophils, neutrophils, lymphocytes, plasma cells), percentage of extracellular Zhou Xieyan cells (eosinophils, neutrophils, lymphocytes).
Bacteria analyzed at the portal level were: proteobacteria, actinomycetes, bacterioides, firmics, fusobacteria.
Bacteria belonging to the horizontal analysis are: staphylococcus, corynebacterium, prevotella, pandorea, bactoides, escherichia/ShigellaClostridium XlVa, moraxella, fusobacterium, streptomyces, achromobacter, propionibacterium, neisseria, dolosporinum, barnesiella, roseburia, faechibacterium, haemophilus peptenophilus, pseudomonas, clostridium sensu stricto, lactobacilli, anaerococcus, veilonella, alloprvoella, bifidobacterium, brevundimonas Porphyromonas, lannospecies_incarnata, capnocytophaga.
Conclusion: clinical indices of differences compared to non-recurrent groups were: 7, respectively: the asthma, olfactory score, symptom score, histoinflammatory cells% (eosinophils, neutrophils, lymphocytes) and peripheral blood eosinophils were pooled.
Compared with a non-recurrent group, the recurrent group has the advantages of increased probability of combined asthma, more serious olfactory disorder and higher symptom total score before operation, higher percentage of tissue and peripheral blood eosinophils, and lower percentage of tissue neutrophils and lymphocytes.
The number of the door level bacterial change indexes is 2, and the indexes are respectively as follows: proteobacteria, actinobactionbacteria. The recurrent group had more Proteobacteria and lower genus level bacteria Actinobacillus.
The number of the bacterial change indexes belonging to the level is 8, and the bacterial change indexes are respectively: prevoltella, fusobacterium, clostridium sensu stricto, veilonella, corynebacterium, staphylococcus, propionibacterium, anaerobiosis. The recurrent group had more Prevotella, fusobacterium, clostridium sensu stricto and Veilonella, less Corynebacterium, staphylococcus, propionibacterium and Anaerococcus.
2. Multiple binary Logistic regression analysis
Differential clinical and flora indices of recurrent and non-recurrent groups were included in Logistic regression analysis based on single factor analysis results. Because the pre-operative symptom score includes an olfactory score that is not an independent variable indicator, the pre-operative symptom score indicator is not included in the multiple Logistic regression analysis. Because there were differences in gate and genus levels, we divided into 2 analysis groups.
Group I inclusion: combining asthma, preoperative olfactory score, tissue and peripheral blood eosinophils, tissue neutrophils, tissue lymphocytes, door horizontal bacteria Proteobacteria and Actinobacillus. Regression analysis was performed using a stepwise forward approach, with the result that only tissue eosinophil% and portal horizontal bacteria actionbacteria were factors that truly could predict recurrence of nasal polyps.
Group II was incorporated into the combination of asthma, preoperative olfactory scores, tissue and peripheral blood eosinophils, tissue neutrophils, tissue lymphocytes, genus Leptobacter, prevolvulella, fusobacterium, clostridium sensu stricto, veilonella, corynebacterium, staphylococcus, propionibacterium and Anaerococcus. Regression analysis was also performed using a stepwise forward approach, with the result that only tissue eosinophil% and the genus level bacteria Corynebacterium are factors that truly predict recurrence of nasal polyps.
TABLE 1 multiple Logistic regression analysis to determine independent influencing factors for predicting nasal polyp recurrence
3. Assessing predictive value of independent predictive recurrence indicators
Subject work characteristic (ROC) curve analysis is an analysis and evaluation of a two-class discrimination effect, and can determine the prediction accuracy, positive prediction rate and cutoff value of potential predictors. While the area under the curve (AUC) represents the prediction accuracy of the potential predictors. The AUC of tissue eosinophil% for predicting nasal polyp recurrence was 0.908, and the AUC of phylum horizontal bacterium actionobacteria, belonging to horizontal bacterium Corynebacterium for predicting polyp recurrence was 0.786 and 0.811, respectively. Furthermore, the combined organization of eosinophil% and the phylum level bacterium actionobacteria or the genus level bacterium Corynebacterium can significantly increase the predictive value of nasal polyp recurrence (auc=0.962, auc=0.981). This means that in this experimental study, the accuracy of predicting nasal polyp recurrence in combination with eosinophil% and actionobacteria or Corynebacterium abundance was 96.2% and 98.1%, respectively.
Table 2 AUC values for predicting polyp recurrence
Note that: 65 samples were tested in total, 31 in the recurrent group and 34 in the non-recurrent group.
While particular embodiments of the present invention have been illustrated and described, it will be appreciated that various other changes and modifications can be made without departing from the spirit and scope of the invention. It is therefore intended to cover in the appended claims all such changes and modifications that are within the scope of this invention.
Claims (5)
1. Use of a flora abundance detector and a reagent for detecting the percentage of tissue eosinophils for the preparation of a nasal polyp prognosis detector, characterized in that the flora comprises bacteria at the phylum level and/or bacteria at the genus level; the horizontal bacteria of the door areActinobacteriaThe method comprises the steps of carrying out a first treatment on the surface of the The genus level bacteria areCorynebacteriumThe method comprises the steps of carrying out a first treatment on the surface of the The nasal polyp is chronic nasal sinusitis with nasal polyp.
2. The use according to claim 1, wherein the detection template of the abundance of the flora is a genome of a microorganism extracted from a middle nasal passage drop; and acquiring a middle nasal meatus falling object by adopting a middle nasal meatus cotton swab.
3. The use of claim 1, wherein the flora abundance detector comprises any one or more of a primer, a probe, a biosensor, and a chip.
4. The use according to claim 3, wherein the flora abundance detector is designed for the V3-V4 region of the bacterial 16S ribosomal RNA gene.
5. The use according to claim 3, wherein the primer is set forth in SEQ ID NO. 1-2.
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Citations (2)
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Non-Patent Citations (8)
| Title |
|---|
| Chronic rhinosinusitis with nasal polyps is characterized by dysbacteriosis of the nasal microbiota;Thanit Chalermwatanachai等;《SCIENTIFIC RePorTs》;20180521;第8卷;文章编号7926 * |
| Comparison of Subtyping Approaches and the Underlying Drivers of Microbial Signatures for Chronic Rhinosinusitis;Kristi Biswas等;《mSphere》;20190228;第4卷(第1期);文章编号e00679-18 * |
| Thanit Chalermwatanachai等.Chronic rhinosinusitis with nasal polyps is characterized by dysbacteriosis of the nasal microbiota.《SCIENTIFIC RePorTs》.2018,第8卷 * |
| The difference in nasal bacterial microbiome diversity between chronic rhinosinusitis patients with polyps and a control population;Weigang Gan等;《Int Forum Allergy Rhinol》;20190205;第9卷;第582-592页 * |
| The microbiology of chronic rhinosinusitis with and without nasal polyps;QINGHUA LIU等;《Acta Oto-Laryngologica》;20141231;第134卷;1251–1258 * |
| The microbiology of chronic rhinosinusitis with and without nasal;Hong‑Zheng Wei等;《European Archives of Oto-Rhino-Laryngology》;20180322 * |
| Weigang Gan等.The difference in nasal bacterial microbiome diversity between chronic rhinosinusitis patients with polyps and a control population.《Int Forum Allergy Rhinol》.2019,第9卷 * |
| 组织嗜酸性粒细胞对鼻窦炎合并鼻息肉患者治疗后病情复发的预测效果;罗伟等;《检验医学与临床》;20170814(第15期) * |
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