The Application of Endoscopic Debridement Combined with Metagenomic Next-Generation Sequencing Technology in Primary Spinal Infections: A Retrospective Study

The Application of Endoscopic Debridement Combined with Metagenomic Next-Generation Sequencing Technology in Primary Spinal Infections: A Retrospective Study | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article The Application of Endoscopic Debridement Combined with Metagenomic Next-Generation Sequencing Technology in Primary Spinal Infections: A Retrospective Study Xiaofei Feng, Jie Cheng, Luyong Jiang, Jiayi Lin, Qingjiang Pang, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4792457/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 25 Feb, 2025 Read the published version in Journal of Orthopaedic Surgery and Research → Version 1 posted 9 You are reading this latest preprint version Abstract Purpose Spinal endoscopy is a novel minimally invasive spinal surgery technique used in recent years to treat various degenerative spinal diseases. Metagenomic next-generation sequencing (mNGS) is a new method for identifying infectious microorganisms in infectious diseases. We aim to evaluate the application effect of combining spinal endoscopy with mNGS in diagnosing and treating spinal infections. Methods The clinical data of 62 patients with suspected spinal infectious diseases admitted from January 2020 to December 2023 were retrospectively analyzed. All patients underwent spinal endoscopy to obtain tissue specimens, histopathological examination, routine bacterial culture, and mNGS sequencing. Describe the pathogenic microbial spectrum of spinal infection, and compare the differences in sensitivity (true positive rate) and specificity (true negative rate) between the two detection methods. White blood cell (WBC) erythrocyte deposition rate (ESR), C-reactive protein (CRP), visual analog scale (VAS), Japanese Orthopaedic Association (JOA) score, Oswestry Disability Index (ODI), and other clinical results were analyzed. Results In 62 cases, mNGS, microbiological culture, serologic testing, and pathologic examination results were obtained. 49 cases of spinal infections and 13 cases of non-spinal infections were finally diagnosed clinically. Among the 49 patients with spinal infections, there were 31 cases of purulent bacterial infections, 8 cases of tuberculosis infections, and 10 cases of infections with unspecified etiological microorganisms. Among the 13 cases of non-spinal infections, there were 3 cases of spinal tumors, 6 cases of Modic changes of the endplates, and 4 cases of endplate fracture. The positive rate of microbial culture was 36.73% (18/49), and the positive rate of the mNGS test was 71.43% (35/49), which was statistically different from each other (P < 0.01). The sensitivity of the mNGS test was 71.43%, and the specificity of the mNGS test was 84.62%. At the 3-month follow-up, WBC, ESR, and CRP levels were normalized. The VAS, JOA score, and ODI of the lower back and legs at each follow-up point after surgery were significantly improved compared with those before surgery, and the difference was statistically significant (P < 0.01). Conclusion Macrogenomic sequencing technology is fast, efficient, and accurate in detecting pathogenic microorganisms, and has high diagnostic value in the diagnosis and treatment of spinal infections. Spinal endoscopic debridement combined with mNGS can achieve good clinical results. Spinal infection Spinal Endoscopy Metagenomic next-generation sequencing Etiological diagnosis Figures Figure 1 Figure 2 1. Introduction Spinal infectious diseases are caused by pathogenic microorganisms that settle in the spinal column area and spread from the affected segment to adjacent segments or tissues [ 1 ] . It often affects the vertebral body, intervertebral disc, or paravertebral tissue, including vertebral osteomyelitis, discitis, and epidural abscess. According to the types of pathogenic microorganisms, infections are classified as specific and nonspecific infections. Specific infections mainly refer to infections such as tuberculosis or non-tuberculous mycobacteria, fungi, Brucella, etc.; nonspecific infections mainly refer to purulent infections caused by G − or G + bacteria, such as Staphylococcus aureus, Escherichia coli, and various streptococci [ 2 – 4 ] . Clinical manifestations, laboratory tests, imaging findings, and histopathological examinations of spinal infections caused by different pathogens lack specificity. Identifying the pathogenic microorganism is crucial for diagnosing the infection [ 5 ] . Therefore, seeking a simpler and more efficient method to obtain the pathogenic microorganism for early diagnosis is necessary. However, traditional detection methods mainly include culture and molecular biology tests. Although a positive culture is considered the gold standard for diagnosis, this method has shortcomings such as long culture periods, low positivity rates, and high false positive rates. Literature reports a blood culture positivity rate of 30–50% for spinal infectious diseases [ 6 – 8 ] . While the positive rate of culture from biopsy specimens of lesions is less than 60% [ 9 – 10 ] . PCR, as a representative of molecular biology detection, has high sensitivity and specificity but requires the pre-designing of primers, only targeting specific pathogens, which is of great clinical significance for the diagnosis of spinal tuberculosis-related infections. Literature reports that traditional methods rely on diagnosing spinal infectious diseases still cannot definitively identify 25%~60% of the infectious pathogens [ 11 – 14 ] . Therefore, there is an urgent need in clinical practice for a broad-spectrum screening detection method for non-tuberculous infectious pathogens. Metagenomic next-generation sequencing technology, as an emerging pathogen detection method, is a new sequencing method relative to first-generation DNA sequencing technology. It can detect various pathogens without bias, making up for the shortcomings of various traditional pathogen detection methods. This technology features fast speed, high accuracy, high positivity rate, and broad coverage of pathogenic bacteria. It has unique advantages in detecting rare pathogens such as mycoplasma, fungi, and brucella. Currently, it has been widely used in the detection of blood, respiratory tract, cerebrospinal fluid, and other specimens, achieving certain effectiveness [ 15 ] . However, there are relatively few clinical reports on the diagnosis of infectious diseases in bone and joints, mainly based on single-center, small-sample research reports. The traditional treatment modalities for spinal infections include mainly non-surgical and open-surgical treatments. Some scholars believe that most intervertebral space infections can be cured by non-surgical treatment through absolute bed rest, strengthening nutrition, and symptomatic treatment with large doses of sensitive antibacterial drugs [ 16 ] . However, non-surgical treatment can lead to long bed rest, easy to prolong the course of the disease, and easy to lead to lumbar instability and pseudoarthrosis in the later stage [ 17 ] . With the advancement of spinal surgery technology and the development of minimally invasive concepts, intervertebral endoscopic has been widely used in the treatment of lumbar spine diseases, and it has a greater advantage in the treatment of spinal infections. This study analyzed the use of metagenomic next-generation sequencing (mNGS) for diagnosing spinal infectious diseases. The goal was to assess mNGS's effectiveness, advantages, and limitations in diagnosing spinal infections. Additionally, the study aimed to identify factors that affect the detection of pathogens in samples to improve the accuracy of results, reduce false positives and false negatives, and evaluate the potential benefits of combining mNGS with endoscopic treatment for primary spinal infections. 2. Materials and methods 2.1 Patients This study is a retrospective research based on the data of inpatients in our hospital. It involves collecting data on all patients suspected of spinal infection who underwent spinal endoscopy sampling and mNGS diagnosis from January 2020 to December 2023. Incorporation criteria: (1) Suspected spinal infectious diseases diagnosed based on clinical manifestations, imaging, and laboratory examinations, (2)Obtained lesion specimens under spinal endoscopy, (3)Perform mNGS testing and routine laboratory inspection. Exclusion criteria: (1) Not completing all routine tests and mNGS, (2) Specimens were significantly contaminated during sampling, transportation, and handling, or the cold chain transportation failed, (3) Incomplete or unclear medical history. According to the above inclusion and exclusion criteria, a total of 62 patients were included in this study. This study was approved by the Ethics Committee of Ningbo No.2 Hospital, and all patients signed informed consent. 2.2 Blood tests Results of the Rose Bengal test, G test, GM test, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP) level, white blood cell (WBC) count, and tuberculosis tests (T-SPOT.TB) were obtained. All blood tests were performed in the laboratory of our hospital. The testing process is shown in Fig. 1 . 2.3 Sample collection and tests All patients obtain blood and tissue specimens from the lesion site. Blood specimens are collected at the hospital, and tests including blood routine, C-reactive protein, erythrocyte sedimentation rate, enzyme-linked immune-spot assay (T-SPOT.TB), detection of glucan content (GM test), and platelet agglutination test are performed for infectious disease laboratory examination. Tissue specimens are obtained directly at the center of the lesion under spinal endoscopy. Fresh samples from all patients are promptly stored in sterile containers and transported to the laboratory on dry ice for metagenomic next-generation sequencing (mNGS). 2.4 Collection of specimens The spinal endoscopic was advanced into the intervertebral space to monitor lesion clearance and end-plate preparation. The nucleus pulposus was completely removed, the pus in and around the vertebral canal was cleaned, and the damaged and hardened bone was cured. The tissues taken during the operation were stored aseptically and submitted for examination(Figure 2 ). 2.5 mNGS The whole process of mNGS testing strictly follows the principle of aseptic operation. The nucleic acid extraction process is carried out in the biosafety cabinet. The operators use the aseptic consumables, reagents, and the gun head with filter elements according to the SOP process. (1) Nucleic acid extraction Sample 200µl of nucleic acid was obtained after pretreatment and nucleic acid extraction, refer to the manual (Nucleic acid extraction kit, Hangzhou Jieyi Biotechnology Co. LTD.) (2) Library preparation (a) Configuration premix solution: fragmentation enzyme 5µl add fragmentation buffer 10µl preparation of premix solution, vortexing for 5s to mix the solution, instantaneous centrifugation for 5s and then placed on ice. (b)After opening the cartridge, add 40µl of purified magnetic beads to test tube No. 1. 1040µl of absolute ethanol to test tube No. 8. 5µl of ligase to test tube No. 10. 30µl of connector to test tube No. 11. 15µl of premix to test tube No. 12, and 35µl of nucleic acid extract to test tube No. 3 to avoid bubbles. The library is obtained after setting up the software running program. (c)Library quantification: Library quantification was quantified by quantitative real-time PCR (KAPA) technology. (d)Computer sequencing: High-throughput sequencing on Illumina Nextseq with 75 bp single-end sequencing with about 20 million sequences generated for each library. (e) Bioinformatics analysis: Bioinformatics analysis performed human genome sequence reference (GRCh38.p13) filtering as described in the literature, with reads of the remaining sequences and reference database (NCBI; ftp://ftp.ncbi.nlm.nih.gov/genomes/ ) to determine species, read count and read count relative abundance. Negative controls were tested in each sequencing, and negative controls used manually cultured 1104 / ml Jurkat tumor cells, excluding environmental bacteria and reagent background bacteria sequences, such as Acinetobacter, Acinetobacter johnsonii, etc. 2.6 Evaluation index The differences between mNGS and conventional bacterial culture on the sensitivity (true positive rate), specificity (true negative rate), false negative rate, and false negative rate. ESR, CRP, WBC, low back, and leg Visual model scores were analyzed at different time points before and after surgery. VAS, Japanese Orthopaedic Association (JOA) score, Oswestry disability index (Oswestry disability index) ODI) at six-month follow-up. 2.7 Statistical analysis Statistical analyses were performed using the SPSS 25.0 (SPSS Corporation, USA) statistical software package. An Independent sample t-test was used for comparison between conventional bacterial culture and mNGS. The count data (specimen acquisition method, antibiotics applied within 4 weeks, and specimen type) were expressed by example (%), and the comparison of positive rate was performed by χ2 test. The comparison of the true positive rate, true negative rate, false positive rate, and false negative rate of different detection methods was performed using the χ2 test. ANOVA with one-way repeated measures data was used to compare VAS, JOA scores, ODI, ESR, and CRP at all postoperative time points. The effect of different factors on the detection of pathogenic microorganisms was analyzed by non-conditional logistic regression. A difference of P < 0.05 indicated statistical significance. 3. Results 3.1 Patient features Among the 62 patients, 37 were males and 25 were females. Age 14–81 years old (49.4 ± 16.2 y). All pathology specimens were obtained by spinal endoscopic surgery. Combined with the history, clinical manifestations, physical examination, laboratory examination, imaging examination, and intraoperative findings. 49 cases of mNGS were clinically diagnosed as spinal infections and 13 cases were non-spinal infections. Of the 49 cases of spinal infection, 32 were treated with antibiotics before obtaining lesion specimens and 17 were treated without antibiotics. ESR (P < 0.001), CRP ( P < 0.001), and WBC count ( P < 0.001) are all significantly higher in the infected group (Table 1 ). There were 4 cases of cervical, 18 cases of thoracic, and 31 cases of lumbar infections. There was no statistical difference in age or gender between the infected and aseptic groups. (Table 2 ) Table 1 Demographic characteristics of cases included in the study Characteristic Aseptic Infected P value Age(y) 52.9 ± 11.6 47.1 ± 13.4 0.58 Gender(M/F) 8/5 29/20 13.69 ESR(mm/h) 23.64 ± 14.27 82.65 ± 19.81 <0.01 CRP(mg/L) 9.24 ± 3.71 56.72 ± 27.92 <0.01 WBC(10 9 /L) 4.36 ± 1.57 8.27 ± 3.19 <0.01 Table 2 Distribution of results from 62 cases of spinal infection Clinical symptom Focal site Focal pain cervical vertebra(4) lumbar vertebra(8) Focal pain and fever thoracic vertebra(9) lumbar vertebra(17) Focal pain and lower limb pain lumbar vertebra(11) 3.2 Diagnostic results of pathogenic microorganisms 3.2.1 Pathology and culture results of pathogenic microorganism Among the 49 cases of spinal infection, there were 31 cases of suppurative bacterial infection, 8 cases of tuberculosis infection, and 10 cases of spinal infection without clear pathogenic microorganisms. There were 13 cases of non-spinal infection, including spinal tumor in 3 cases, endplate Modic change in 6 cases, and endplate fracture in 4 cases. Among the 49 patients with clear evidence of pathogenic microorganisms, 37 cases were identical with the laboratory results and 12 cases were different from the final diagnosis. The consistency between mNGS and laboratory results was 71.43% (35/49). In the test results of mNGS, the common pathogenic microorganisms of suppurative spondylitis were Staphylococcus aureus, Escherichia coli, streptococcus, and fungi. The other common infection type was spinal tuberculosis. 7 cases of tuberculosis infection were detected by pathology and quantitative PCR, and the smear and culture were negative (Table 3 ). Table 3 Comparison of clinically confirmed pathogen microorganism results with metagenomic sequencing results. Patient ID Pathogenic microorganisms Diagnostic Basis Results of mNGS and sequence number Antibiotic 1–11 Stapyhlococcus aureus Bacterial culture Stapyhlococcus aureus Penicillin 12–14 MRSA Bacterial culture Stapyhlococcus aureus Vancomycin 15–16 Streptococcus sanguis Bacterial culture,PCR Prohyromonas endodontalis, Treponema deticola, Streptococcus sanguis Penicillin Metronidazole 17 Streptococcus intermedium Bacterial culture Streptococcus intermedium Linezolid, Minocycline 18–19 Streptococcus constellations Bacterial culture Streptococcus constellations Linezolid, Minocycline 20–34 Escherichia coil Bacterial culture Escherichia coil Cephalosporin, Levofloxacin 25–27 Pseidomonas aeruginosa Blood culture Pseidomonas aeruginosa Cephalosporin, Levofloxacin 28–29 Bacteroides fragilis Blood culture Bacteroides fragilis Carbapenems, Metronidazole 30 Aspergillus flavus Bacterial culture Aspergillus flavus Metronidazole 31–39 Mycobacterium tuberculosis Pathology, PCR Mycobacterium Tuberculosis complex Rifampicin, Isoniazid, Ethambutol, Pyrazinamide 40 Unknown pathogen Pathology Streptococcus pseudopneumoniae Rifampicin, Isoniazid, Ethambutol, Pyrazinamide 41 Unknown pathogen Pathology Escherichia coli Vancomycin༎Carbapenems༎ 42 Unknown pathogen Pathology Streptococcus sinensis Cephalosporin, Levofloxacin 43 Unknown pathogen Pathology Staphylococcus epidermidis Linezolid, Levofloxacin 44–46 Unknown pathogen Pathology Staphylococcus aureus Linezolid, Levofloxacin 47 Unknown pathogen Pathology Negative Vancomycin 48 Unknown pathogen Pathology Negative Cephalosporin, Levofloxacin 49 Unknown pathogen Pathology Negative Carbapenems 50–52 Spinal metastasis Pathology Negative N/A 53–58 Modic changes Pathology Negative N/A 59–62 Endplate fracture Pathology Negative N/A 3.2.2 mNGS test results The differences in sensitivity (true positive rate), specificity (true negative rate), false positive rate, and false negative rate of pathogenic microorganisms detection between mNGS and conventional bacterial culture were compared. The study suggests that the differences in sensitivity and false-negative rate between mNGS and conventional bacterial culture are statistically significant, indicating that mNGS can effectively improve the detection rate of pathogenic microorganisms (Table 4 ). Table 4 Specificity and sensitivity of mNGS compared with culture Test result bacterial culture mNGS χ 2 P sensitivity 36.73% 71.43% 12.73 < 0.001 false negative rate 63.27% 28.57% < 0.001 false positive rate 46.15% 15.38% < 0.001 specificity 53.85% 84.62% 11.64 < 0.001 3.3 Comparison of Evaluation Index at 1 year of follow-up The patients had immediate relief of lower back pain after surgery. All patients were followed up for 12 to 18 months, with a mean of 14 months. ESR, CRP, and WBC were all reduced to the normal range at 1 month postoperatively. The difference between the preoperative VAS, JOA score, and ODI index on the first day, first month, third month, sixth month, and the last follow-up was statistically significant (P < 0.05)(Table 5 ). Table 5 The VAS scores of low back and Low limb pain, JOA score, ODI, ESR, CRP, and WBC were compared between preoperative and postoperative. Test Point VAS(Low back pain) VAS(Low limb pain) JOA (score) ODI (%) ESR (m/h) CRP (mg/L) WBC (10 9 /L) preoperative 6.34 ± 1.72 4.07 ± 1.14 12.82 ± 4.68 61.36 ± 9.32 97.52 ± 25.48 65.72 ± 29.15 9.67 ± 3.58 Postoperative Day 3.32 ± 0.63 1.87 ± 1.09 - - 89.15 ± 19.76 63.18 ± 24.76 9.55 ± 4.24 One month after the operative 1.89 ± 0.43 1.32 ± 0.76 21.57 ± 2.89 36.26 ± 8.93 20.17 ± 10.48 8.46 ± 4.27 3.83 ± 1.62 Three months after the operative 1.37 ± 1.65 1.06 ± 0.53 25.28 ± 2.47 27.48 ± 5.95 19.71 ± 8.84 7.98 ± 3.47 3.86 ± 1.06 Six months after the operative 0.79 ± 0.58 0.71 ± 0.47 26.27 ± 3.73 9.85 ± 4.63 21.08 ± 10.67 8.68 ± 4.28 4.02 ± 1.34 One year after the operative 0.25 ± 0.37 0.28 ± 0.24 28.83 ± 4.36 7.92 ± 3.45 18.63 ± 9.76 9.27 ± 3.89 3.87 ± 1.06 P < 0.001 < 0.001 < 0.001 < 0.001 < 0.001 < 0.001 < 0.001 Discussion The diagnosis of spinal infections can be challenging. Initially, spinal infections share clinical and imaging similarities with spinal tumors, seronegative spondylitis fractures, and endplate injuries in the early stages, making them difficult to distinguish. Secondly, spinal infections are mainly caused by pyogenic bacteria, Mycobacterium brucei, Mycobacterium tuberculosis, and fungi. The clinical features, signs, and imaging manifestations often lack specificity, making them easily confused. The accurate and efficient diagnosis of spinal infections and other infectious diseases has always relied on high-quality diagnostic techniques. While microbiologic culture is considered the gold standard for diagnosing spinal infections, the positive rate of bacterial culture for Mycobacterium brucei, Mycobacterium tuberculosis, fungi, and anaerobes is extremely low. Serologic testing is a specific test for one or more microorganisms with unique properties. While highly targeted, it is subject to cross-reactivity, changes in the course of the disease, and the influence of background potency, which may result in false positives or false negatives. Xu reported an overall positive rate of 90.72% for mNGS in 108 cases of spinal infections, which was significantly higher than the 40% for routine cultures [ 18 ] . Wang found that the sensitivity of mNGS in the detection of spinal infection pathogens using one- and two-generation high-throughput sequencing techniques was 73.33% [ 19 ] . Ma reported that the sensitivity of mNGS in 31 patients with spinal infections was 70.3% and the specificity was 75.0% [ 20 ] . In the present study, the sensitivity and specificity of the mNGS assay reached 71.43% and 84.62%, respectively. Similar to the results of other studies, mNGS was significantly superior to a combination of culture, serologic testing, pathology, and PCR assays. We assumed that the differences in the diagnostic ability of mNGS in different studies are mainly due to the differences in sequencing platforms, testing centers, and sample tissues. mNGS is not culture-dependent, and its detection results only depend on the DNA content of the pathogenic microorganisms at the time of DNA library establishment. For microorganisms that are difficult to isolate DNA from, such as Mycobacterium tuberculosis, Mycobacterium brucei, and other intracellular bacteria or fungi, the optimization of the extraction process and interpretation rules can be used to improve the detection rate. In addition, even if the pathogenic microorganisms die after anti-infection treatment, their DNA remains active for a short period. However, their DNA can retain activity in the short term, and results can still be obtained by mNGS detection. In this study, some patients were still found to have pathogenic microorganisms by mNGS after anti-infective treatment, indicating that mNGS is less affected by anti-infective treatment in the short term than other detection methods. DNA extraction, library construction, sequencing, and analysis of pathogenic microorganisms in all patients in this study were all completed within 48 hours. The process was fully completed within 48h, which is comparable to serological testing and shorter than conventional culture time (3, 7d, and some cultures even need to be extended to 14d). mNGS's high efficiency and rapidity imply early intervention of the disease and thus reduce the risk of exacerbation. Another advantage of mNGS is that it is comprehensive and low-biased. The range of mNGS results covers 6350 species of bacteria, 1798 DNA viruses, 1064 fungi, and 234 species with known genome sequences. Bacteria, 1798 DNA viruses, 1064 fungi, and 234 parasites. Parasites, essentially including all spinal infection pathogenic microorganisms. Although the difficulty of DNA extraction from different species, the bias of the test compared with the conventional test. However, the bias is low compared with conventional tests. Theoretically, it is possible to directly obtain all the genetic information in the specimen. It excludes the subjective purpose and intervention of the detector, which makes up for the tendency and limitation of various single-laboratory tests. And limitations of various single-laboratory tests. At the same time, different from alveolar lavage fluid, pus, and other specimens of infectious diseases, the presence of unclear foci and localized contaminants affect the interpretation of mNGS results, the spine is a naturally sterile environment. The spine is a natural sterile environment, and the foci are relatively clear and confined after the occurrence of infection. Therefore, when mNGS detects rare and uncommon pathogenic microorganisms, it usually has higher accuracy and confidence. In this study, mNGS detection identified multiple infections, anaerobic infections, specific types of infections, and other rare microbial infections were identified by mNGS, and the diagnosis was confirmed clinically by combining history, imaging, special laboratory tests, and diagnostic treatment. Compared with spinal puncture biopsy, spinal endoscopic lesion removal and sampling can provide enough tissue for smear tests, microbial culture using various culture media, and pathological examination. This ensures a comprehensive clinical testing program, reducing the risk of missed diagnosis and misdiagnosis. While taking samples, we also removed the lesion. During the operation, we cleared out pus, pathogenic bacteria, inflammatory factors, and necrotic tissues by using large amounts of saline for rinsing. As a result, the patient's postoperative back and leg pain was immediately relieved. Conclusion Spinal endoscopic debridement combined with metagenomic next-generation sequencing (mNGS) for the diagnosis and treatment of primary spinal infections combines the advantages of endoscopic surgery for direct lesion removal and mNGS for precise and rapid diagnosis, which can achieve favorable clinical outcomes. Declarations Ethics approval and consent to participate Patients' informed consent was obtained (Ethics Committee of Ningbo No. 2 Hospital, Ref. No. YJ-NBEY-KYSB-2024-100-12). Consent for publication Not applicable. Availability of data and materials Data will be made available on request. Competing interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Funding This research was supported by the project of Ningbo leading medical & Health Discipline(2022-F15), the Key Research Foundation of Ningbo No.2 Hospital (2024HMZD05), the Distinguished Young Scholars of Ningbo No.2 Hospital(2023HMJQ03). Authors' contributions Xiaofei Feng: Conceptualization, Methodology, Formal analysis, Investigation, Data curation, Writing. Jie Cheng: Methodology, Data curation, Validation. Luyong Jiang: Methodology, Validation. Jiayi Lin: Methodology, Investigation. Zhewei Ye: Methodology, Resources. Qingjiang Pang: Supervision, Writing. Jiangtao: Supervision, Project administration, Funding acquisition. Acknowledgments We thank all patients for providing consent to our use of cases. We thank all financial support. Thanks for the technical support by the Medical Laboratory department of Ningbo No.2 Hospital. References Luzzati R, Giacomazzi D, Danzi MC, et al. Diagnosis, management, and outcome of clinically- suspected spinal infection. 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Cite Share Download PDF Status: Published Journal Publication published 25 Feb, 2025 Read the published version in Journal of Orthopaedic Surgery and Research → Version 1 posted Editorial decision: Revision requested 23 Sep, 2024 Reviews received at journal 23 Sep, 2024 Reviews received at journal 21 Sep, 2024 Reviewers agreed at journal 06 Sep, 2024 Reviewers agreed at journal 05 Sep, 2024 Reviewers invited by journal 03 Sep, 2024 Editor assigned by journal 24 Jul, 2024 Submission checks completed at journal 24 Jul, 2024 First submitted to journal 24 Jul, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4792457","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":334968916,"identity":"881b4c45-351a-487f-923a-b7d91fd9b465","order_by":0,"name":"Xiaofei Feng","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA4ElEQVRIie3PMQrCMBSA4ZRAuiTtJimF9gopXT1Mg+DUwdGxoMRFnRUv4ej4SqFT9AIughcQXEQLWhwcbUbBfMMjw/t5BCHL+kXgnNrZp8SdAFzGRgkW7RxGHq1ludLmSZVGPE8rNjUovGPlnEd3LBXVF2AFiv0efE+CwwCn6yWRyl1sIdihZL3JvidC+3XI5rS9st9ColEmjp0Jdh9szqXi+QmkMksIpjeREp4jKE2SQGMcsiKLCK1FWWje/RdPY+dKmyeNZ5PztRn3Yz/sSN4c9Xlyg/W3xnTRsizrL70AytJJGVcQXb4AAAAASUVORK5CYII=","orcid":"","institution":"Ningbo No. 2 Hospital","correspondingAuthor":true,"prefix":"","firstName":"Xiaofei","middleName":"","lastName":"Feng","suffix":""},{"id":334968917,"identity":"599c264d-8bdd-424a-a1b1-8b26c63b21f2","order_by":1,"name":"Jie Cheng","email":"","orcid":"","institution":"Wenzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Jie","middleName":"","lastName":"Cheng","suffix":""},{"id":334968918,"identity":"ecfbbcb6-6621-443a-88eb-1f99412f3036","order_by":2,"name":"Luyong Jiang","email":"","orcid":"","institution":"Ningbo No. 2 Hospital","correspondingAuthor":false,"prefix":"","firstName":"Luyong","middleName":"","lastName":"Jiang","suffix":""},{"id":334968919,"identity":"e4e7c2e3-5922-4bcf-8762-13d868bf57bb","order_by":3,"name":"Jiayi Lin","email":"","orcid":"","institution":"Ningbo No. 2 Hospital","correspondingAuthor":false,"prefix":"","firstName":"Jiayi","middleName":"","lastName":"Lin","suffix":""},{"id":334968920,"identity":"bf6b1ea9-83a3-4055-8760-20002f8137e7","order_by":4,"name":"Qingjiang Pang","email":"","orcid":"","institution":"Ningbo No. 2 Hospital","correspondingAuthor":false,"prefix":"","firstName":"Qingjiang","middleName":"","lastName":"Pang","suffix":""},{"id":334968921,"identity":"3197efcc-356d-4f7e-b8f2-43cbb19cff0e","order_by":5,"name":"Jiangtao Liu","email":"","orcid":"","institution":"Ningbo No. 2 Hospital","correspondingAuthor":false,"prefix":"","firstName":"Jiangtao","middleName":"","lastName":"Liu","suffix":""},{"id":334968922,"identity":"b8ed6c30-1ab5-413a-bfbc-7d9c816c2fe4","order_by":6,"name":"Zhewei Ye","email":"","orcid":"","institution":"Huazhong University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Zhewei","middleName":"","lastName":"Ye","suffix":""}],"badges":[],"createdAt":"2024-07-24 05:11:31","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4792457/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4792457/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s13018-024-05385-5","type":"published","date":"2025-02-25T15:58:15+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":63298624,"identity":"2c7afc61-d90b-4e17-8eb4-f3565aa7ef44","added_by":"auto","created_at":"2024-08-26 15:48:23","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":156603,"visible":true,"origin":"","legend":"\u003cp\u003eLaboratory examination and aging chart of patients with spinal infection\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-4792457/v1/2e282b38094a2fd02168f566.png"},{"id":63298625,"identity":"bc5cb977-5328-42db-9307-fede7856b692","added_by":"auto","created_at":"2024-08-26 15:48:23","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":774639,"visible":true,"origin":"","legend":"\u003cp\u003eThe focal specimens were collected under a spinal endoscope.\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-4792457/v1/bf4f7d876cbf321e1885f30c.png"},{"id":77623224,"identity":"ac5a3981-a562-411a-9574-adaf563b533c","added_by":"auto","created_at":"2025-03-03 16:11:15","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2250728,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4792457/v1/b7cdb640-ef11-4c86-8879-d951042d3058.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"The Application of Endoscopic Debridement Combined with Metagenomic Next-Generation Sequencing Technology in Primary Spinal Infections: A Retrospective Study","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eSpinal infectious diseases are caused by pathogenic microorganisms that settle in the spinal column area and spread from the affected segment to adjacent segments or tissues\u003csup\u003e[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]\u003c/sup\u003e. It often affects the vertebral body, intervertebral disc, or paravertebral tissue, including vertebral osteomyelitis, discitis, and epidural abscess. According to the types of pathogenic microorganisms, infections are classified as specific and nonspecific infections. Specific infections mainly refer to infections such as tuberculosis or non-tuberculous mycobacteria, fungi, Brucella, etc.; nonspecific infections mainly refer to purulent infections caused by G\u003csup\u003e\u0026minus;\u003c/sup\u003e or G\u003csup\u003e+\u003c/sup\u003e bacteria, such as Staphylococcus aureus, Escherichia coli, and various streptococci\u003csup\u003e[\u003cspan additionalcitationids=\"CR3\" citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eClinical manifestations, laboratory tests, imaging findings, and histopathological examinations of spinal infections caused by different pathogens lack specificity. Identifying the pathogenic microorganism is crucial for diagnosing the infection\u003csup\u003e[\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]\u003c/sup\u003e. Therefore, seeking a simpler and more efficient method to obtain the pathogenic microorganism for early diagnosis is necessary. However, traditional detection methods mainly include culture and molecular biology tests. Although a positive culture is considered the gold standard for diagnosis, this method has shortcomings such as long culture periods, low positivity rates, and high false positive rates. Literature reports a blood culture positivity rate of 30\u0026ndash;50% for spinal infectious diseases\u003csup\u003e[\u003cspan additionalcitationids=\"CR7\" citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]\u003c/sup\u003e. While the positive rate of culture from biopsy specimens of lesions is less than 60%\u003csup\u003e[\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]\u003c/sup\u003e. PCR, as a representative of molecular biology detection, has high sensitivity and specificity but requires the pre-designing of primers, only targeting specific pathogens, which is of great clinical significance for the diagnosis of spinal tuberculosis-related infections. Literature reports that traditional methods rely on diagnosing spinal infectious diseases still cannot definitively identify 25%~60% of the infectious pathogens\u003csup\u003e[\u003cspan additionalcitationids=\"CR12 CR13\" citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e14\u003c/span\u003e]\u003c/sup\u003e. Therefore, there is an urgent need in clinical practice for a broad-spectrum screening detection method for non-tuberculous infectious pathogens.\u003c/p\u003e \u003cp\u003eMetagenomic next-generation sequencing technology, as an emerging pathogen detection method, is a new sequencing method relative to first-generation DNA sequencing technology. It can detect various pathogens without bias, making up for the shortcomings of various traditional pathogen detection methods. This technology features fast speed, high accuracy, high positivity rate, and broad coverage of pathogenic bacteria. It has unique advantages in detecting rare pathogens such as mycoplasma, fungi, and brucella. Currently, it has been widely used in the detection of blood, respiratory tract, cerebrospinal fluid, and other specimens, achieving certain effectiveness\u003csup\u003e[\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e15\u003c/span\u003e]\u003c/sup\u003e. However, there are relatively few clinical reports on the diagnosis of infectious diseases in bone and joints, mainly based on single-center, small-sample research reports.\u003c/p\u003e \u003cp\u003eThe traditional treatment modalities for spinal infections include mainly non-surgical and open-surgical treatments. Some scholars believe that most intervertebral space infections can be cured by non-surgical treatment through absolute bed rest, strengthening nutrition, and symptomatic treatment with large doses of sensitive antibacterial drugs\u003csup\u003e[\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e16\u003c/span\u003e]\u003c/sup\u003e. However, non-surgical treatment can lead to long bed rest, easy to prolong the course of the disease, and easy to lead to lumbar instability and pseudoarthrosis in the later stage\u003csup\u003e[\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e17\u003c/span\u003e]\u003c/sup\u003e. With the advancement of spinal surgery technology and the development of minimally invasive concepts, intervertebral endoscopic has been widely used in the treatment of lumbar spine diseases, and it has a greater advantage in the treatment of spinal infections.\u003c/p\u003e \u003cp\u003eThis study analyzed the use of metagenomic next-generation sequencing (mNGS) for diagnosing spinal infectious diseases. The goal was to assess mNGS's effectiveness, advantages, and limitations in diagnosing spinal infections. Additionally, the study aimed to identify factors that affect the detection of pathogens in samples to improve the accuracy of results, reduce false positives and false negatives, and evaluate the potential benefits of combining mNGS with endoscopic treatment for primary spinal infections.\u003c/p\u003e"},{"header":"2. Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\n \u003ch2\u003e2.1 Patients\u003c/h2\u003e\n \u003cp\u003eThis study is a retrospective research based on the data of inpatients in our hospital. It involves collecting data on all patients suspected of spinal infection who underwent spinal endoscopy sampling and mNGS diagnosis from January 2020 to December 2023. Incorporation criteria: (1) Suspected spinal infectious diseases diagnosed based on clinical manifestations, imaging, and laboratory examinations, (2)Obtained lesion specimens under spinal endoscopy, (3)Perform mNGS testing and routine laboratory inspection. Exclusion criteria: (1) Not completing all routine tests and mNGS, (2) Specimens were significantly contaminated during sampling, transportation, and handling, or the cold chain transportation failed, (3) Incomplete or unclear medical history. According to the above inclusion and exclusion criteria, a total of 62 patients were included in this study. This study was approved by the Ethics Committee of Ningbo No.2 Hospital, and all patients signed informed consent.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec4\" class=\"Section2\"\u003e\n \u003ch2\u003e2.2 Blood tests\u003c/h2\u003e\n \u003cp\u003eResults of the Rose Bengal test, G test, GM test, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP) level, white blood cell (WBC) count, and tuberculosis tests (T-SPOT.TB) were obtained. All blood tests were performed in the laboratory of our hospital. The testing process is shown in Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e\n \u003ch2\u003e2.3 Sample collection and tests\u003c/h2\u003e\n \u003cp\u003eAll patients obtain blood and tissue specimens from the lesion site. Blood specimens are collected at the hospital, and tests including blood routine, C-reactive protein, erythrocyte sedimentation rate, enzyme-linked immune-spot assay (T-SPOT.TB), detection of glucan content (GM test), and platelet agglutination test are performed for infectious disease laboratory examination. Tissue specimens are obtained directly at the center of the lesion under spinal endoscopy. Fresh samples from all patients are promptly stored in sterile containers and transported to the laboratory on dry ice for metagenomic next-generation sequencing (mNGS).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec6\" class=\"Section2\"\u003e\n \u003ch2\u003e2.4 Collection of specimens\u003c/h2\u003e\n \u003cp\u003eThe spinal endoscopic was advanced into the intervertebral space to monitor lesion clearance and end-plate preparation. The nucleus pulposus was completely removed, the pus in and around the vertebral canal was cleaned, and the damaged and hardened bone was cured. The tissues taken during the operation were stored aseptically and submitted for examination(Figure \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e\n \u003ch2\u003e2.5 mNGS\u003c/h2\u003e\n \u003cp\u003eThe whole process of mNGS testing strictly follows the principle of aseptic operation. The nucleic acid extraction process is carried out in the biosafety cabinet. The operators use the aseptic consumables, reagents, and the gun head with filter elements according to the SOP process.\u003c/p\u003e\u003cspan\u003e\n \u003cp\u003e\u003cstrong\u003e(1) Nucleic acid extraction\u003c/strong\u003e\u003c/p\u003e\n \u003c/span\u003e\n \u003cp\u003eSample 200\u0026micro;l of nucleic acid was obtained after pretreatment and nucleic acid extraction, refer to the manual (Nucleic acid extraction kit, Hangzhou Jieyi Biotechnology Co. LTD.)\u003c/p\u003e\u003cspan\u003e\n \u003cp\u003e\u003cstrong\u003e(2) Library preparation\u003c/strong\u003e\u003c/p\u003e\n \u003c/span\u003e\u003cspan\u003e\n \u003cp\u003e(a) Configuration premix solution: fragmentation enzyme 5\u0026micro;l add fragmentation buffer 10\u0026micro;l preparation of premix solution, vortexing for 5s to mix the solution, instantaneous centrifugation for 5s and then placed on ice. (b)After opening the cartridge, add 40\u0026micro;l of purified magnetic beads to test tube No. 1. 1040\u0026micro;l of absolute ethanol to test tube No. 8. 5\u0026micro;l of ligase to test tube No. 10. 30\u0026micro;l of connector to test tube No. 11. 15\u0026micro;l of premix to test tube No. 12, and 35\u0026micro;l of nucleic acid extract to test tube No. 3 to avoid bubbles. The library is obtained after setting up the software running program. (c)Library quantification: Library quantification was quantified by quantitative real-time PCR (KAPA) technology. (d)Computer sequencing: High-throughput sequencing on Illumina Nextseq with 75 bp single-end sequencing with about 20 million sequences generated for each library. (e) Bioinformatics analysis: Bioinformatics analysis performed human genome sequence reference (GRCh38.p13) filtering as described in the literature, with reads of the remaining sequences and reference database (NCBI; \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003eftp://ftp.ncbi.nlm.nih.gov/genomes/\u003c/span\u003e\u003c/span\u003e) to determine species, read count and read count relative abundance. Negative controls were tested in each sequencing, and negative controls used manually cultured 1104 / ml Jurkat tumor cells, excluding environmental bacteria and reagent background bacteria sequences, such as Acinetobacter, Acinetobacter johnsonii, etc.\u003c/p\u003e\n \u003c/span\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\n \u003ch2\u003e2.6 Evaluation index\u003c/h2\u003e\n \u003cp\u003eThe differences between mNGS and conventional bacterial culture on the sensitivity (true positive rate), specificity (true negative rate), false negative rate, and false negative rate. ESR, CRP, WBC, low back, and leg Visual model scores were analyzed at different time points before and after surgery. VAS, Japanese Orthopaedic Association (JOA) score, Oswestry disability index (Oswestry disability index) ODI) at six-month follow-up.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\n \u003ch2\u003e2.7 Statistical analysis\u003c/h2\u003e\n \u003cp\u003eStatistical analyses were performed using the SPSS 25.0 (SPSS Corporation, USA) statistical software package. An Independent sample t-test was used for comparison between conventional bacterial culture and mNGS. The count data (specimen acquisition method, antibiotics applied within 4 weeks, and specimen type) were expressed by example (%), and the comparison of positive rate was performed by \u0026chi;2 test. The comparison of the true positive rate, true negative rate, false positive rate, and false negative rate of different detection methods was performed using the \u0026chi;2 test. ANOVA with one-way repeated measures data was used to compare VAS, JOA scores, ODI, ESR, and CRP at all postoperative time points. The effect of different factors on the detection of pathogenic microorganisms was analyzed by non-conditional logistic regression. A difference of P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 indicated statistical significance.\u003c/p\u003e\n\u003c/div\u003e"},{"header":"3. Results","content":"\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003e3.1 Patient features\u003c/h2\u003e \u003cp\u003eAmong the 62 patients, 37 were males and 25 were females. Age 14–81 years old (49.4 ± 16.2 y). All pathology specimens were obtained by spinal endoscopic surgery. Combined with the history, clinical manifestations, physical examination, laboratory examination, imaging examination, and intraoperative findings. 49 cases of mNGS were clinically diagnosed as spinal infections and 13 cases were non-spinal infections. Of the 49 cases of spinal infection, 32 were treated with antibiotics before obtaining lesion specimens and 17 were treated without antibiotics. ESR (P \u0026lt; 0.001), CRP (\u003cem\u003eP\u003c/em\u003e \u0026lt; 0.001), and WBC count (\u003cem\u003eP\u003c/em\u003e \u0026lt; 0.001) are all significantly higher in the infected group (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). There were 4 cases of cervical, 18 cases of thoracic, and 31 cases of lumbar infections. There was no statistical difference in age or gender between the infected and aseptic groups. (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e)\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eDemographic characteristics of cases included in the study\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCharacteristic\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAseptic\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eInfected\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eP value\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAge(y)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e52.9 ± 11.6\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e47.1 ± 13.4\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.58\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGender(M/F)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8/5\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e29/20\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e13.69\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eESR(mm/h)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e23.64 ± 14.27\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e82.65 ± 19.81\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;0.01\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCRP(mg/L)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9.24 ± 3.71\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e56.72 ± 27.92\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;0.01\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eWBC(10\u003csup\u003e9\u003c/sup\u003e/L)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4.36 ± 1.57\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e8.27 ± 3.19\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt;0.01\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eDistribution of results from 62 cases of spinal infection\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"2\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eClinical symptom\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eFocal site\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFocal pain\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ecervical vertebra(4) lumbar vertebra(8)\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFocal pain and fever\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ethoracic vertebra(9) lumbar vertebra(17)\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFocal pain and lower limb pain\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003elumbar vertebra(11)\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e3.2 Diagnostic results of pathogenic microorganisms\u003c/h2\u003e \u003cdiv id=\"Sec13\" class=\"Section3\"\u003e \u003ch2\u003e3.2.1 Pathology and culture results of pathogenic microorganism\u003c/h2\u003e \u003cp\u003eAmong the 49 cases of spinal infection, there were 31 cases of suppurative bacterial infection, 8 cases of tuberculosis infection, and 10 cases of spinal infection without clear pathogenic microorganisms. There were 13 cases of non-spinal infection, including spinal tumor in 3 cases, endplate Modic change in 6 cases, and endplate fracture in 4 cases. Among the 49 patients with clear evidence of pathogenic microorganisms, 37 cases were identical with the laboratory results and 12 cases were different from the final diagnosis. The consistency between mNGS and laboratory results was 71.43% (35/49). In the test results of mNGS, the common pathogenic microorganisms of suppurative spondylitis were Staphylococcus aureus, Escherichia coli, streptococcus, and fungi. The other common infection type was spinal tuberculosis. 7 cases of tuberculosis infection were detected by pathology and quantitative PCR, and the smear and culture were negative (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComparison of clinically confirmed pathogen microorganism results with metagenomic sequencing results.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePatient ID\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePathogenic microorganisms\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eDiagnostic\u003c/p\u003e \u003cp\u003eBasis\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eResults of mNGS and sequence number\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eAntibiotic\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1–11\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eStapyhlococcus\u003c/p\u003e \u003cp\u003eaureus\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacterial culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStapyhlococcus aureus\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ePenicillin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e12–14\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eMRSA\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacterial culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStapyhlococcus aureus\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eVancomycin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e15–16\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eStreptococcus sanguis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacterial culture,PCR\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eProhyromonas endodontalis, Treponema deticola,\u003c/p\u003e \u003cp\u003eStreptococcus sanguis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003ePenicillin\u003c/p\u003e \u003cp\u003eMetronidazole\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e17\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eStreptococcus intermedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacterial culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStreptococcus intermedium\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eLinezolid, Minocycline\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e18–19\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eStreptococcus constellations\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacterial culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStreptococcus constellations\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eLinezolid, Minocycline\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e20–34\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eEscherichia coil\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacterial culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eEscherichia coil\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eCephalosporin, Levofloxacin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e25–27\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePseidomonas aeruginosa\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBlood culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePseidomonas aeruginosa\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eCephalosporin, Levofloxacin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e28–29\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBacteroides fragilis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBlood culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eBacteroides fragilis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eCarbapenems, Metronidazole\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e30\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAspergillus flavus\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacterial culture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eAspergillus flavus\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eMetronidazole\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e31–39\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eMycobacterium\u003c/p\u003e \u003cp\u003etuberculosis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology,\u003c/p\u003e \u003cp\u003ePCR\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eMycobacterium\u003c/p\u003e \u003cp\u003eTuberculosis complex\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eRifampicin, Isoniazid, Ethambutol, Pyrazinamide\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e40\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStreptococcus pseudopneumoniae\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eRifampicin, Isoniazid, Ethambutol, Pyrazinamide\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e41\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eEscherichia coli\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eVancomycin༎Carbapenems༎\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e42\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStreptococcus sinensis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eCephalosporin, Levofloxacin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e43\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStaphylococcus epidermidis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eLinezolid, Levofloxacin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e44–46\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStaphylococcus aureus\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eLinezolid, Levofloxacin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e47\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNegative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eVancomycin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e48\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNegative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eCephalosporin, Levofloxacin\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e49\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eUnknown pathogen\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNegative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eCarbapenems\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e50–52\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSpinal metastasis\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNegative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eN/A\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e53–58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eModic changes\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNegative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eN/A\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e59–62\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eEndplate fracture\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePathology\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNegative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eN/A\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section3\"\u003e \u003ch2\u003e3.2.2 mNGS test results\u003c/h2\u003e \u003cp\u003eThe differences in sensitivity (true positive rate), specificity (true negative rate), false positive rate, and false negative rate of pathogenic microorganisms detection between mNGS and conventional bacterial culture were compared. The study suggests that the differences in sensitivity and false-negative rate between mNGS and conventional bacterial culture are statistically significant, indicating that mNGS can effectively improve the detection rate of pathogenic microorganisms (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSpecificity and sensitivity of mNGS compared with culture\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTest result\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ebacterial culture\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003emNGS\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eχ\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eP\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003esensitivity\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e36.73%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e71.43%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e12.73\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003efalse negative rate\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e63.27%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e28.57%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003efalse positive rate\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e46.15%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e15.38%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003especificity\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e53.85%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e84.62%\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e11.64\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e3.3 Comparison of Evaluation Index at 1 year of follow-up\u003c/h2\u003e \u003cp\u003eThe patients had immediate relief of lower back pain after surgery. All patients were followed up for 12 to 18 months, with a mean of 14 months. ESR, CRP, and WBC were all reduced to the normal range at 1 month postoperatively. The difference between the preoperative VAS, JOA score, and ODI index on the first day, first month, third month, sixth month, and the last follow-up was statistically significant (P \u0026lt; 0.05)(Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe VAS scores of low back and Low limb pain, JOA score, ODI, ESR, CRP, and WBC were compared between preoperative and postoperative.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"8\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTest Point\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eVAS(Low back pain)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eVAS(Low limb pain)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eJOA\u003c/p\u003e \u003cp\u003e(score)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eODI\u003c/p\u003e \u003cp\u003e(%)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eESR\u003c/p\u003e \u003cp\u003e(m/h)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eCRP\u003c/p\u003e \u003cp\u003e(mg/L)\u003c/p\u003e \u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eWBC\u003c/p\u003e \u003cp\u003e(10\u003csup\u003e9\u003c/sup\u003e/L)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003epreoperative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.34 ± 1.72\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4.07 ± 1.14\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e12.82 ± 4.68\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e61.36\u003c/p\u003e \u003cp\u003e± 9.32\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e97.52\u003c/p\u003e \u003cp\u003e± 25.48\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e65.72\u003c/p\u003e \u003cp\u003e± 29.15\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e9.67\u003c/p\u003e \u003cp\u003e± 3.58\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePostoperative Day\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3.32 ± 0.63\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.87 ± 1.09\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e89.15\u003c/p\u003e \u003cp\u003e± 19.76\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e63.18\u003c/p\u003e \u003cp\u003e± 24.76\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e9.55\u003c/p\u003e \u003cp\u003e± 4.24\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eOne month after the operative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1.89 ± 0.43\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.32 ± 0.76\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e21.57 ± 2.89\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e36.26\u003c/p\u003e \u003cp\u003e± 8.93\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e20.17\u003c/p\u003e \u003cp\u003e± 10.48\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e8.46\u003c/p\u003e \u003cp\u003e± 4.27\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e3.83\u003c/p\u003e \u003cp\u003e± 1.62\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eThree months after the operative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1.37 ± 1.65\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.06 ± 0.53\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e25.28 ± 2.47\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e27.48\u003c/p\u003e \u003cp\u003e± 5.95\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e19.71\u003c/p\u003e \u003cp\u003e± 8.84\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e7.98\u003c/p\u003e \u003cp\u003e± 3.47\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e3.86\u003c/p\u003e \u003cp\u003e± 1.06\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSix months after the operative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.79 ± 0.58\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.71 ± 0.47\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e26.27 ± 3.73\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e9.85\u003c/p\u003e \u003cp\u003e± 4.63\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e21.08\u003c/p\u003e \u003cp\u003e± 10.67\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e8.68\u003c/p\u003e \u003cp\u003e± 4.28\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e4.02\u003c/p\u003e \u003cp\u003e± 1.34\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eOne year after the operative\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.25 ± 0.37\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.28 ± 0.24\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e28.83 ± 4.36\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e7.92\u003c/p\u003e \u003cp\u003e± 3.45\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e18.63\u003c/p\u003e \u003cp\u003e± 9.76\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e9.27\u003c/p\u003e \u003cp\u003e± 3.89\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e3.87\u003c/p\u003e \u003cp\u003e± 1.06\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt; 0.001\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e \u003cp\u003e\u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe diagnosis of spinal infections can be challenging. Initially, spinal infections share clinical and imaging similarities with spinal tumors, seronegative spondylitis fractures, and endplate injuries in the early stages, making them difficult to distinguish. Secondly, spinal infections are mainly caused by pyogenic bacteria, Mycobacterium brucei, Mycobacterium tuberculosis, and fungi. The clinical features, signs, and imaging manifestations often lack specificity, making them easily confused. The accurate and efficient diagnosis of spinal infections and other infectious diseases has always relied on high-quality diagnostic techniques. While microbiologic culture is considered the gold standard for diagnosing spinal infections, the positive rate of bacterial culture for Mycobacterium brucei, Mycobacterium tuberculosis, fungi, and anaerobes is extremely low. Serologic testing is a specific test for one or more microorganisms with unique properties. While highly targeted, it is subject to cross-reactivity, changes in the course of the disease, and the influence of background potency, which may result in false positives or false negatives.\u003c/p\u003e\u003cp\u003eXu reported an overall positive rate of 90.72% for mNGS in 108 cases of spinal infections, which was significantly higher than the 40% for routine cultures\u003csup\u003e[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e18\u003c/span\u003e]\u003c/sup\u003e. Wang found that the sensitivity of mNGS in the detection of spinal infection pathogens using one- and two-generation high-throughput sequencing techniques was 73.33% \u003csup\u003e[\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e19\u003c/span\u003e]\u003c/sup\u003e. Ma reported that the sensitivity of mNGS in 31 patients with spinal infections was 70.3% and the specificity was 75.0%\u003csup\u003e[\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e20\u003c/span\u003e]\u003c/sup\u003e. In the present study, the sensitivity and specificity of the mNGS assay reached 71.43% and 84.62%, respectively. Similar to the results of other studies, mNGS was significantly superior to a combination of culture, serologic testing, pathology, and PCR assays.\u003c/p\u003e\u003cp\u003eWe assumed that the differences in the diagnostic ability of mNGS in different studies are mainly due to the differences in sequencing platforms, testing centers, and sample tissues. mNGS is not culture-dependent, and its detection results only depend on the DNA content of the pathogenic microorganisms at the time of DNA library establishment. For microorganisms that are difficult to isolate DNA from, such as Mycobacterium tuberculosis, Mycobacterium brucei, and other intracellular bacteria or fungi, the optimization of the extraction process and interpretation rules can be used to improve the detection rate. In addition, even if the pathogenic microorganisms die after anti-infection treatment, their DNA remains active for a short period. However, their DNA can retain activity in the short term, and results can still be obtained by mNGS detection. In this study, some patients were still found to have pathogenic microorganisms by mNGS after anti-infective treatment, indicating that mNGS is less affected by anti-infective treatment in the short term than other detection methods. DNA extraction, library construction, sequencing, and analysis of pathogenic microorganisms in all patients in this study were all completed within 48 hours. The process was fully completed within 48h, which is comparable to serological testing and shorter than conventional culture time (3, 7d, and some cultures even need to be extended to 14d). mNGS's high efficiency and rapidity imply early intervention of the disease and thus reduce the risk of exacerbation.\u003c/p\u003e\u003cp\u003eAnother advantage of mNGS is that it is comprehensive and low-biased. The range of mNGS results covers 6350 species of bacteria, 1798 DNA viruses, 1064 fungi, and 234 species with known genome sequences. Bacteria, 1798 DNA viruses, 1064 fungi, and 234 parasites. Parasites, essentially including all spinal infection pathogenic microorganisms. Although the difficulty of DNA extraction from different species, the bias of the test compared with the conventional test. However, the bias is low compared with conventional tests. Theoretically, it is possible to directly obtain all the genetic information in the specimen. It excludes the subjective purpose and intervention of the detector, which makes up for the tendency and limitation of various single-laboratory tests. And limitations of various single-laboratory tests. At the same time, different from alveolar lavage fluid, pus, and other specimens of infectious diseases, the presence of unclear foci and localized contaminants affect the interpretation of mNGS results, the spine is a naturally sterile environment. The spine is a natural sterile environment, and the foci are relatively clear and confined after the occurrence of infection. Therefore, when mNGS detects rare and uncommon pathogenic microorganisms, it usually has higher accuracy and confidence. In this study, mNGS detection identified multiple infections, anaerobic infections, specific types of infections, and other rare microbial infections were identified by mNGS, and the diagnosis was confirmed clinically by combining history, imaging, special laboratory tests, and diagnostic treatment.\u003c/p\u003e\u003cp\u003eCompared with spinal puncture biopsy, spinal endoscopic lesion removal and sampling can provide enough tissue for smear tests, microbial culture using various culture media, and pathological examination. This ensures a comprehensive clinical testing program, reducing the risk of missed diagnosis and misdiagnosis. While taking samples, we also removed the lesion. During the operation, we cleared out pus, pathogenic bacteria, inflammatory factors, and necrotic tissues by using large amounts of saline for rinsing. As a result, the patient's postoperative back and leg pain was immediately relieved.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eSpinal endoscopic debridement combined with metagenomic next-generation sequencing (mNGS) for the diagnosis and treatment of primary spinal infections combines the advantages of endoscopic surgery for direct lesion removal and mNGS for precise and rapid diagnosis, which can achieve favorable clinical outcomes.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePatients\u0026apos; informed consent was obtained (Ethics Committee of Ningbo No. 2 Hospital, Ref. No. YJ-NBEY-KYSB-2024-100-12).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData will be made available on request.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research was supported by the project of Ningbo leading medical \u0026amp; Health Discipline(2022-F15), the Key Research Foundation of Ningbo No.2 Hospital\u003c/p\u003e\n\u003cp\u003e(2024HMZD05), the Distinguished Young Scholars of Ningbo No.2 Hospital(2023HMJQ03).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026apos; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eXiaofei Feng: Conceptualization, Methodology, Formal analysis, Investigation, Data curation, Writing. Jie Cheng: Methodology, Data curation, Validation. Luyong Jiang: \u0026nbsp; Methodology, Validation. Jiayi Lin: Methodology, Investigation. Zhewei Ye: Methodology, Resources. Qingjiang Pang: Supervision, Writing. Jiangtao: Supervision, Project administration, Funding acquisition.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank all patients for providing consent to our use of cases. We thank all financial support. Thanks for the technical support by the Medical Laboratory department of Ningbo No.2 Hospital.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eLuzzati R, Giacomazzi D, Danzi MC, et al. Diagnosis, management, and outcome of clinically- suspected spinal infection. J Infect. 2009 Apr;58(4):259-65. doi: 10.1016/j.jinf.2009.02.006.\u003c/li\u003e\n \u003cli\u003eKenyon PC, Chapman AL. Tuberculous vertebral osteomyelitis: findings of a 10-year review of experience in a UK centre. J Infect. 2009 Nov;59(5):372-3. doi: 10.1016/j.jinf.2009.09.011.\u003c/li\u003e\n \u003cli\u003eDuarte RM, Vaccaro AR. Spinal infection: state of the art and management algorithm. Eur Spine J. 2013 Dec;22(12):2787-99. doi: 10.1007/s00586-013-2850-1.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eHuang H, Shi J, Zheng M, et al. Pathogen detection in suspected spinal infection: metagenomic next-generation sequencing versus culture. Eur Spine J. 2023 Dec;32(12):4220-4228. doi: 10.1007/s00586-023-07707-3.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eKim CJ, Song KH, Park WB, et al. Microbiologically and clinically diagnosed vertebral osteomyelitis: impact of prior antibiotic exposure. Antimicrob Agents Chemother. 2012 Apr;56(4):2122-4. doi: 10.1128/AAC.05953-11.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eOverbeek R, Leitl CJ, Stoll SE, et al. The Value of Next-Generation Sequencing in Diagnosis and Therapy of Critically Ill Patients with Suspected Bloodstream Infections: A Retrospective Cohort Study. J Clin Med. 2024 Jan 5;13(2):306. doi: 10.3390/jcm13020306.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eBurillo A, Bouza E. Use of rapid diagnostic techniques in ICU patients with infections. BMC Infect Dis. 2014 Nov 28;14:593. doi: 10.1186/s12879-014-0593-1.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eWu S, Hu W, Xiao W, et al. Metagenomic Next-Generation Sequencing Assists in the Diagnosis of Gardnerella vaginalis in Males with Pleural Effusion and Lung Infection: A Case Report and Literature Review. Infect Drug Resist. 2021 Dec 7;14:5253-5259. doi: 10.2147/IDR.S337248.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eSehn JK, Gilula LA. Percutaneous needle biopsy in diagnosis and identification of causative organisms in cases of suspected vertebral osteomyelitis. Eur J Radiol. 2012 May;81(5):940-6. doi: 10.1016/j.ejrad.2011.01.125.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eTerreaux W, Geoffroy M, Ohl X, et al. Diagnostic contribution of a second percutaneous needle biopsy in patients with spontaneous diskitis and negative blood cultures and first biopsy. Joint Bone Spine. 2016 Dec;83(6):715-719. doi: 10.1016/j.jbspin.2016.02.006.\u003c/li\u003e\n \u003cli\u003eHoupikian P, Raoult D. Blood culture-negative endocarditis in a reference center: etiologic diagnosis of 348 cases. Medicine (Baltimore). 2005 May;84(3):162-173. doi 10.1097/01.md.0000165658.82869.17.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003ePallen MJ. Diagnostic metagenomics: potential applications to bacterial, viral and parasitic infections. Parasitology. 2014 Dec;141(14):1856-62. doi:10.1017/S00311820 14000134.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eGeissd\u0026ouml;rfer W, Moos V, Moter A, et al. High frequency of Tropheryma whipplei in culture-negative endocarditis. J Clin Microbiol. 2012 Feb;50(2):216-22. doi: 10.1128/JCM.05531-11.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eOechslin CP, Lenz N, Liechti N, et al. Limited Correlation of Shotgun Metagenomics Following Host Depletion and Routine Diagnostics for Viruses and Bacteria in Low Concentrated Surrogate and Clinical Samples. Front Cell Infect Microbiol. 2018 Oct 23;8:375. doi: 10.3389/fcimb.2018.00375.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eGu W, Miller S, Chiu CY. Clinical Metagenomic Next-Generation Sequencing for Pathogen Detection. Annu Rev Pathol. 2019 Jan 24;14:319-338. doi: 10.1146/annurev-pathmechdis-012418-012751.\u003c/li\u003e\n \u003cli\u003eWalters R, Vernon-Roberts B, Fraser R, Moore R. Therapeutic use of cephazolin to prevent complications of spine surgery. Inflammopharmacology. 2006 Aug;14(3-4):138-43. doi: 10.1007/s10787-006-1503-y.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eBas T, Bas P, Blasco A, Bas JL. Chronic infections of the spine. Eur J Orthop Surg Traumatol. 2013 Jul;23 Suppl 1:S35-40. doi: 10.1007/s00590-013-1245-7.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eXu L, Zhou Z, Wang Y, Song C, Tan H. Improved accuracy of etiological diagnosis of spinal infection by metagenomic next-generation sequencing. Front Cell Infect Microbiol. 2022 Oct 7;12:929701. doi: 10.3389/fcimb.2022.929701.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eWang G, Long J, Zhuang Y, et al. Application of metagenomic next-generation sequencing in the detection of pathogens in spinal infections. Spine J. 2023 Jun;23(6):859-867. doi: 10.1016/j.spinee.2023.02.001.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eMa C, Wu H, Chen G, et al. The potential of metagenomic next-generation sequencing in diagnosis of spinal infection: a retrospective study. Eur Spine J. 2022 Feb;31(2):442-447. doi: 10.1007/s00586-021-07026-5.\u0026nbsp;\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-orthopaedic-surgery-and-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"josr","sideBox":"Learn more about [Journal of Orthopaedic Surgery and Research](http://josr-online.biomedcentral.com)","snPcode":"13018","submissionUrl":"https://submission.nature.com/new-submission/13018/3","title":"Journal of Orthopaedic Surgery and Research","twitterHandle":"@MSKmedBMC","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Spinal infection, Spinal Endoscopy, Metagenomic next-generation sequencing, Etiological diagnosis","lastPublishedDoi":"10.21203/rs.3.rs-4792457/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4792457/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003ePurpose\u003c/h2\u003e \u003cp\u003eSpinal endoscopy is a novel minimally invasive spinal surgery technique used in recent years to treat various degenerative spinal diseases. Metagenomic next-generation sequencing (mNGS) is a new method for identifying infectious microorganisms in infectious diseases. We aim to evaluate the application effect of combining spinal endoscopy with mNGS in diagnosing and treating spinal infections.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eThe clinical data of 62 patients with suspected spinal infectious diseases admitted from January 2020 to December 2023 were retrospectively analyzed. All patients underwent spinal endoscopy to obtain tissue specimens, histopathological examination, routine bacterial culture, and mNGS sequencing. Describe the pathogenic microbial spectrum of spinal infection, and compare the differences in sensitivity (true positive rate) and specificity (true negative rate) between the two detection methods. White blood cell (WBC) erythrocyte deposition rate (ESR), C-reactive protein (CRP), visual analog scale (VAS), Japanese Orthopaedic Association (JOA) score, Oswestry Disability Index (ODI), and other clinical results were analyzed.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eIn 62 cases, mNGS, microbiological culture, serologic testing, and pathologic examination results were obtained. 49 cases of spinal infections and 13 cases of non-spinal infections were finally diagnosed clinically. Among the 49 patients with spinal infections, there were 31 cases of purulent bacterial infections, 8 cases of tuberculosis infections, and 10 cases of infections with unspecified etiological microorganisms. Among the 13 cases of non-spinal infections, there were 3 cases of spinal tumors, 6 cases of Modic changes of the endplates, and 4 cases of endplate fracture. The positive rate of microbial culture was 36.73% (18/49), and the positive rate of the mNGS test was 71.43% (35/49), which was statistically different from each other (P\u0026thinsp;\u0026lt;\u0026thinsp;0.01). The sensitivity of the mNGS test was 71.43%, and the specificity of the mNGS test was 84.62%. At the 3-month follow-up, WBC, ESR, and CRP levels were normalized. The VAS, JOA score, and ODI of the lower back and legs at each follow-up point after surgery were significantly improved compared with those before surgery, and the difference was statistically significant (P\u0026thinsp;\u0026lt;\u0026thinsp;0.01).\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eMacrogenomic sequencing technology is fast, efficient, and accurate in detecting pathogenic microorganisms, and has high diagnostic value in the diagnosis and treatment of spinal infections. Spinal endoscopic debridement combined with mNGS can achieve good clinical results.\u003c/p\u003e","manuscriptTitle":"The Application of Endoscopic Debridement Combined with Metagenomic Next-Generation Sequencing Technology in Primary Spinal Infections: A Retrospective Study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-08-26 15:48:18","doi":"10.21203/rs.3.rs-4792457/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-09-24T01:30:48+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-09-24T01:14:03+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-09-21T08:30:09+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"226885269699778639971992950556035055040","date":"2024-09-07T01:46:44+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"72011354803521463180020420453190365436","date":"2024-09-05T07:12:34+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-09-04T01:27:52+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-07-25T00:58:21+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-07-25T00:26:14+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Orthopaedic Surgery and Research","date":"2024-07-24T05:10:11+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-orthopaedic-surgery-and-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"josr","sideBox":"Learn more about [Journal of Orthopaedic Surgery and Research](http://josr-online.biomedcentral.com)","snPcode":"13018","submissionUrl":"https://submission.nature.com/new-submission/13018/3","title":"Journal of Orthopaedic Surgery and Research","twitterHandle":"@MSKmedBMC","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"28c8153c-ac41-4d52-a2ec-b6242fbcfc95","owner":[],"postedDate":"August 26th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-03-03T16:08:40+00:00","versionOfRecord":{"articleIdentity":"rs-4792457","link":"https://doi.org/10.1186/s13018-024-05385-5","journal":{"identity":"journal-of-orthopaedic-surgery-and-research","isVorOnly":false,"title":"Journal of Orthopaedic Surgery and Research"},"publishedOn":"2025-02-25 15:58:15","publishedOnDateReadable":"February 25th, 2025"},"versionCreatedAt":"2024-08-26 15:48:18","video":"","vorDoi":"10.1186/s13018-024-05385-5","vorDoiUrl":"https://doi.org/10.1186/s13018-024-05385-5","workflowStages":[]},"version":"v1","identity":"rs-4792457","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4792457","identity":"rs-4792457","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}