Generation Sequencing Technologies Research Articles

Next Generation Sequencing Technologies for the analysis of a poorly investigated foodstuff: mushroom products authentication by metabarcoding

Multi-species mushroom-based products (MPs) sold in Italy were authenticated by metabarcoding (MB). One degenerated primer pair was projected for the ITS-1 region amplification. The final data were interpreted using positive controls and extraction blanks. Sequences of macrofungi, microfungi and bacteria were detected in 100%, 90.9% and 77.3% of the MPs, respectively. Not edible and toxic macrofungi were also found, although in low sequence amount. The match between species declared on MPs label and species detected by MB was 59.1%. Cases of voluntary species substitution cannot be excluded. Methods to authenticate these products, still poorly investigated, should be further employed, and MB protocol should be standardized to be used in the context of both official control and companies’ self-control. Alternative approaches should be considered for canned and frozen MPs, in virtue of the observed DNA fragmentation.

Identifying gut microbiota with high specificity for ischemic stroke with large vessel occlusion

Gut microbiota can regulate the metabolic and immunological aspects of ischemic stroke and modulate the treatment effects. The present study aimed to identify specific changes in gut microbiota in patients with large vessel occlusion (LVO) ischemic stroke and assess the potential association between gut microbiota and clinical features of ischemic stroke. A total of 63 CSVD patients, 64 cerebral small vessel disease (CSVD) patients, and 36 matching normal controls (NCs) were included in this study. The fecal samples were collected for all participants and analyzed for gut microbiota using 16S rRNA gene sequencing technology. The abundances of five gut microbiota, including genera Bifidobacterium, Butyricimonas, Blautia, and Dorea and species Bifidobacterium_longum, showed significant changes with high specificity in the LVO patients as compared to the NCs and CSVD patients. In LVO patients, the genera Bifidobacterium and Blautia and species Bifidobacterium_longum were significantly correlated with the National Institutes of Health Stroke Scale (NIHSS) scores at the admission and discharge of the patients. Serum triglyceride levels could significantly affect the association of the abundance of genus Bifidobacterium and species Bifidobacterium_longum with the NIHSS scores at admission and modified Rankin Scale (mRS) at discharge in LVO patients. The identification of five gut microbiota with high specificity were identified in the early stage of LVO stroke, which contributed to performed an effective clinical management for LVO ischemic stroke.

An Ensemble Cascade Forest‐Based Framework for Multi‐Omics Drug Response and Synergy Prediction

The obscure drug response continues to be a limiting factor for accurate cures for cancer. Next generation sequencing technologies have propelled the pharmacogenomic studies with characterized large panels of cancer cell line at multi‐omics level. However, the sufficient integration of the multi‐omics data and the efficient prediction for drug response and synergy still remain a challenge. To address these problems, ECFD is designed, an ensemble cascade forest‐based framework that predicts drug response and synergy using five types of omics data. Experimental results show the significant advantages of the ECFD model over existing models. The best integration of feature extraction is determined and the superiorities of robust stability in the face of new and small samples are highlighted. In addition, the methodological framework highlights the explainability of the model, the mechanisms of drug resistance and drug combination treatment strategies based on explainable analyses and biological networks. In sum, ECFD may facilitate the evaluation of drug response and speculation of potential synergy therapies in personalized and precision treatment.

Genomic profiling of lung adenocarcinoma and squamous carcinoma in Indian patients: Implications for precision oncology.

e20599 Background: Lung cancer, the leading cause of cancer related mortalities, remains a significant global health burden. Despite advances in next generation sequencing technologies, the genomic landscape of non-small cell lung cancer (NSCLC) in the Indian patients remains underexplored. In this study, we aimed to elucidate the genomic landscape of lung adenocarcinoma and squamous carcinoma among Indian patients using targeted sequencing data of over 400 lung cancer patients. Methods: Utilizing targeted sequencing of 50 established cancer-related genes, we conducted a comprehensive analysis of the genomic alteration profiles in 325 lung adenocarcinoma and 81 lung squamous carcinoma samples from Indian patients. Further, we employed robust statistical methodologies to delineate correlations between genomic alterations and clinical characteristics including age, gender, extent of disease, habits and PD-L1 status. Furthermore, our investigation revealed distinctive genomic signatures of NSCLC specific to the Indian population through comparative analyses across various ethnicities. Results: EGFR mutations were detected in 45.8% of lung adenocarcinoma and a notable 18% of lung squamous carcinoma cases among Indian patients, showing significant prevalence among females and non-smokers. Alongside EGFR, lung adenocarcinoma patients exhibited frequent alterations in TP53 (27.4%), ALK (11.4%), and KRAS (10.2%), while lung squamous carcinoma cases predominantly showed mutations in TP53 (40.7%), PIK3CA (17.3%), and CDKN2A (8.6%). Remarkably, 68% of lung adenocarcinoma cases deemed eligible for NCCN-approved targeted therapies. Additionally, it was observed that 60% of PD-L1-negative squamous carcinoma patients harbored TP53 alterations, indicating intriguing therapeutic implications. Overall, we observed alterations in 88% lung adenocarcinoma and 70% lung squamous carcinoma patients. Conclusions: Our study unveiled distinct genomic variations present in both adenocarcinoma and squamous carcinoma patients from India, revealing crucial insights for precision medicine and clinical management of lung cancer. Our findings provide a significant lead for designing clinical trials aimed at evaluating EGFR inhibitors in lung squamous carcinoma patients, potentially expanding therapeutic options for Indian lung cancer patients.

Prevalence of NTRK gene fusions in solid tumors in northeastern Mexico.

e15145 Background: Gene fusions (GF) are among the intricate hallmarks of cancer that lead to abnormal cell proliferation and survival. Neurotrophic tropomyosin receptor kinase (NTRK) gene fusions are frequent in rare solid tumors but less frequent in common ones. Scarce literature has evaluated the prevalence of this aberration estimated to be from 0.5 to 1% globally. For these particular type of tumors, multikinase inhibitors is an option aside from chemotherapy with good responses. Objective: To determine the prevalence of NTRK gene fusions in a Mexican population. Methods: Sociodemographic, clinical, and histopathological data were collected in a single center from databases of Pathology and Oncology Departments of Hospital Universitario Dr. José E. González in Northeastern Mexico. Patients of all ages with solid tumors were included if tested positive for Pan-TRK by immunohistochemistry (IHC). Confirmation for gene fusion was made using Next Generation Sequencing (NGS) technology in formalin-fixed paraffin-embedded tissue. Results: A total of 156 patients from 2021 to 2023 were included for analysis. 67 (42.9%) did test positive for IHC. Of these, 52.2% were male and 20.8% were pediatric. Adult and pediatric median age of 57 and 13 years, respectively. Most frequent tumors locations were central nervous system & extremities. Gene fusion anomaly was confirmed in 6 patients. Among these, 83.3% were male and 16.7% pediatric population. Only NTRK1 and NTRK3 were found in gene fusions. Four translocations were prevalent TPM3:NTRK1 and ETV6:NTRK3 with 2 cases (33.3%), respectively. Followed by LMNA:NTRK1 and EML4:NTRK3 each with one patient. For location, arranged in descending order, head and neck (33.3%), extremities (33.3%), central nervous system and occult primary tumors both 16.6%. The only non-adult tumor reported was infantile fibrosarcoma. Prevalence was estimated to be 3.8% in our population. Conclusions: Understanding the molecular biology of tumor cells enables the development of targeted treatments other than chemotherapy. In our population, NTRK gene fusions were a little more prevalent (~4%) than global estimation. Unfortunately, in Mexico there's a lack of access to multikinase inhibitors. Further studies with large samples are required in Mexico and all over the world to afford access to these tumor-agnostic therapies.

Variant Detection in 3′ Exons of PMS2 Using Exome Sequencing Data

PMS2 is one of the mismatch repair genes included in routine genetic testing for Lynch syndrome, colorectal, ovarian, and endometrial cancers. PMS2 is also included in the American College of Medical Genetics and Genomics (ACMG) secondary findings gene list in the context of clinical exome and genome sequencing. However, sequencing of PMS2 by short-read based next generation sequencing (NGS) technologies is complicated by the presence of the pseudogene PMS2CL and often supplemented by long-range based approaches such as long-range polymerase chain reaction (LR-PCR) or long-read based next generation sequencing, which increases the complexity and cost. Here, we described a bioinformatics homology triage workflow that can eliminate the need for long-read based testing for PMS2 for the vast majority of patients undergoing exome sequencing, thus simplifying PMS2 testing and reducing the associated cost.

Structure-based network analysis predicts pathogenic variants in human proteins associated with inherited retinal disease.

Advances in gene sequencing technologies have accelerated the identification of genetic variants, but better tools are needed to understand which are causal of disease. This would be particularly useful in fields where gene therapy is a potential therapeutic modality for a disease-causing variant such as inherited retinal disease (IRD). Here, we apply structure-based network analysis (SBNA), which has been successfully utilized to identify variant-constrained amino acid residues in viral proteins, to identify residues that may cause IRD if subject to missense mutation. SBNA is based entirely on structural first principles and is not fit to specific outcome data, which makes it distinct from other contemporary missense prediction tools. In 4 well-studied human disease-associated proteins (BRCA1, HRAS, PTEN, and ERK2) with high-quality structural data, we find that SBNA scores correlate strongly with deep mutagenesis data. When applied to 47 IRD genes with available high-quality crystal structure data, SBNA scores reliably identified disease-causing variants according to phenotype definitions from the ClinVar database. Finally, we applied this approach to 63 patients at Massachusetts Eye and Ear (MEE) with IRD but for whom no genetic cause had been identified. Untrained models built using SBNA scores and BLOSUM62 scores for IRD-associated genes successfully predicted the pathogenicity of novel variants (AUC = 0.851), allowing us to identify likely causative disease variants in 40 IRD patients. Model performance was further augmented by incorporating orthogonal data from EVE scores (AUC = 0.927), which are based on evolutionary multiple sequence alignments. In conclusion, SBNA can used to successfully identify variants as causal of disease in human proteins and may help predict variants causative of IRD in an unbiased fashion.

Molecular diagnosis of hen eggs microbiota.

The study was conducted in the Kingdom of Saudi Arabia from 2020 and 2022. The identification, characterization, and evaluation of microbes found in hen eggs was done and it was found very important to prevent contamination caused by various harmful pathogenic microbes. It was found that contaminated eggs harbor various harmful microbes which affect health due to multiple infectious diseases. Hen eggs contain a wide variety of microbes, and several distinct approaches were utilized as well as available for achieving detailed pathogenic information. The information obtained is highly essential for people who consume eggs as a food product. It is of the utmost importance to protect people from getting sick due to the consumption of contaminated eggs or eggs from chickens that have been infected by various harmful pathogens. During the experiment, we found that eggs were contaminated directly or the chicken that laid the egg was contaminated. Using molecular genetic analysis, it is possible to detect pathogenic and non-pathogenic contaminations in eggs. During present studies, the cutting-edge molecular techniques of 16S rRNA gene sequencing technology were used to carry out the objective of performing a molecular identification of the microbial communities infecting eggs. The present research is aimed at determining whether the microbial communities in hen eggs are harmful to humans. The results further indicated most bacteria have the potential to cause illness in humans including Escherichia fergusonii, Salmonella enterica, Pseudocitrobacter faecalis, Yakenella regensburgei, and Erwinia pyrifoliae. Further, research suggested that eggs need to be properly cooked and thoroughly washed to eliminate the possibility of consuming infected eggs.

Illuminating mitochondrial translation through mouse models.

Mitochondria are hubs of metabolic activity with a major role in ATP conversion by oxidative phosphorylation (OXPHOS). The mammalian mitochondrial genome encodes 11 mRNAs encoding 13 OXPHOS proteins along with 2 rRNAs and 22 tRNAs, that facilitate their translation on mitoribosomes. Maintaining the internal production of core OXPHOS subunits requires modulation of the mitochondrial capacity to match the cellular requirements and correct insertion of particularly hydrophobic proteins into the inner mitochondrial membrane. The mitochondrial translation system is essential for energy production and defects result in severe, phenotypically diverse diseases, including mitochondrial diseases that typically affect postmitotic tissues with high metabolic demands. Understanding the complex mechanisms that underlie the pathologies of diseases involving impaired mitochondrial translation is key to tailoring specific treatments and effectively targeting the affected organs. Disease mutations have provided a fundamental, yet limited, understanding of mitochondrial protein synthesis, since effective modification of the mitochondrial genome has proven challenging. However, advances in next generation sequencing, cryoelectron microscopy, and multi-omic technologies have revealed unexpected and unusual features of the mitochondrial protein synthesis machinery in the last decade. Genome editing tools have generated unique models that have accelerated our mechanistic understanding of mitochondrial translation and its physiological importance. Here we review the most recent mouse models of disease pathogenesis caused by defects in mitochondrial protein synthesis and discuss their value for preclinical research and therapeutic development.

A varying-coefficient model for the analysis of methylation sequencing data

DNA methylation is an important epigenetic modification involved in gene regulation. Advances in the next generation sequencing technology have enabled the retrieval of DNA methylation information at single-base-resolution. However, due to the sequencing process and the limited amount of isolated DNA, DNA-methylation-data are often noisy and sparse, which complicates the identification of differentially methylated regions (DMRs), especially when few replicates are available. We present a varying-coefficient model for detecting DMRs by using single-base-resolved methylation information. The model simultaneously smooths the methylation profiles and allows detection of DMRs, while accounting for additional covariates. The proposed model takes into account possible overdispersion by using a beta-binomial distribution. The overdispersion itself can be modeled as a function of the genomic region and explanatory variables. We illustrate the properties of the proposed model by applying it to two real-life case studies.

Research progress on ultrastructure of Treponema pallidum subspecies pallidum

The ultrastructure of Treponema pallidum, the causative pathogen of syphilis, remains unclarified because of the difficulty of long-term culture in vitro. However, research on the ultrastructure of T. pallidum has made great progress in recent years with the success of long-term culture of T. pallidum in vitro and the development of gene sequencing technology and electron microscopy. This review summarizes the research progress on the ultrastructure of T. pallidum.

The effects of inulin on solubilizing and improving anti-obesity activity of high polymerization persimmon tannin

High polymerization persimmon tannin has been reported to have lipid-lowering effects. Unfortunately, the poor solubility restricts its application. This research aimed to investigate the effect and mechanism of inulin on solubilizing of persimmon tannin. Furthermore, we examined whether the addition of inulin would affect the attenuated obesity effect of persimmon tannin. Transmission electron microscope (TEM), Isothermal titration calorimetry (ITC) and Fourier transform infrared spectroscopy (FT-IR) results demonstrated that inulin formed a gel-like network structure, which enabled the encapsulation of persimmon tannin through hydrophobic and hydrogen bond interactions, thereby inhibiting the self-aggregation of persimmon tannin. The turbidity of the persimmon tannin solution decreased by 56.2 %, while the polyphenol content in the supernatant increased by 60.0 %. Furthermore, biochemical analysis and 16s rRNA gene sequencing technology demonstrated that persimmon tannin had a significant anti-obesity effect and improved intestinal health in HFD-fed mice. Moreover, inulin was found to have a positive effect on enhancing the health benefits of persimmon tannin, including improving hepatic steatosis and gut microbiota dysbiosis. it enhanced the abundance of beneficial core microbes while decreasing the abundance of harmful bacteria. Our findings expand the applications of persimmon tannin in the food and medical sectors.

The preliminary application of mNGS in the diagnosis of invasive fungal sinusitis

Objective: By conducting a retrospective analysis of the clinical data of 14 patients diagnosed with invasive fungal rhinosinusitis (IFRS) confirmed by metagenomics next generation sequencing (mNGS) technology, we aim to explore the rapid diagnosis value of mNGS in IFRS. Methods: The clinical data of 14 IFRS patients admitted to TianJin First Central Hospital were retrospectively analyzed from February 2021 to October 2023. The study cohort comprised 8 males and 6 females, with ages ranging from 14 to 77 years. All patients were diagnosed as IFRS by performing mNGS sequencing technology of nasal sinus lesion biopsy specimens. Clinical data such as laboratory examination, imaging examination, histopathological examination results, treatment plan and prognosis were summarized and analyzed. Results: All 14 patients were diagnosed as IFRS, with mNGS detecting pathogens such as Rhizopus (7 cases), Aspergillus (5 cases), Trichoderma (1 case), and Scedosporium apiospermum (1 case). Follow-up evaluations were conducted for a period ranging from 2 months to 2 years post-treatment. At the end of follow-up, 11 out of 14 IFRS patients achieved a complete cure with no signs of recurrence, while the symptoms of the remaining 3 patients significantly improved with comprehensive treatment. Conclusion: mNGS emerges as a highly effective diagnostic tool for IFRS, providing valuable microbiological evidence for clinical diagnosis and demonstrating promising clinical utility.

Impact of high-altitude acclimatization and de-acclimatization on the intestinal microbiota of rats in a natural high-altitude environment.

Intestinal microorganisms play an important role in the health of both humans and animals, with their composition being influenced by changes in the host's environment. We evaluated the longitudinal changes in the fecal microbial community of rats at different altitudes across various time points. Rats were airlifted to high altitude (3,650 m) and acclimatized for 42 days (HAC), before being by airlifted back to low altitude (500 m) and de-acclimatized for 28 days (HADA); meanwhile, the control group included rats living at low altitude (500 m; LA). We investigated changes in the gut microbiota at 12 time points during high-altitude acclimatization and de-acclimatization, employing 16S rRNA gene sequencing technology alongside physiological indices, such as weight and daily autonomous activity time. A significant increase in the Chao1 index was observed on day 14 in the HAC and HADA groups compared to that in the LA group, indicating clear differences in species richness. Moreover, the principal coordinate analysis revealed that the bacterial community structures of HAC and HADA differed from those in LA. Long-term high-altitude acclimatization and de- acclimatization resulted in the reduced abundance of the probiotic Lactobacillus. Altitude and age significantly influenced intestinal microbiota composition, with changes in ambient oxygen content and atmospheric partial pressure being considered key causal factors of altitude-dependent alterations in microbiota composition. High-altitude may be linked to an increase in anaerobic bacterial abundance and a decrease in non-anaerobic bacterial abundance. In this study, the hypobaric hypoxic conditions at high-altitude increased the abundance of anaerobes, while reducing the abundance of probiotics; these changes in bacterial community structure may, ultimately, affect host health. Overall, gaining a comprehensive understanding of the intestinal microbiota alterations during high-altitude acclimatization and de-acclimatization is essential for the development of effective prevention and treatment strategies to better protect the health of individuals traveling between high- and low-altitude areas.

Response of sediment microbial communities to the flow effect of the triangular artificial reef: A simulation-based experimental study

Artificial reefs (ARs), as an important tool for habitat restoration, play significant impacts on benthic microbial ecosystems. This study utilized 16S rRNA gene sequencing technology and computational fluid dynamics (CFD) flow simulation to investigate the effects of flow field distribution around ARs on microbial community structure. The results revealed distinct regional distribution patterns of microbial communities affected by different hydrodynamic conditions. Flow velocity and flow regime of water in sediment-water interface shaped the microbial community structure. The diversity and richness in R–HF were significantly decreased compared to other five regions (p < 0.05). At the phyla and OUT levels, most abundant taxa (1>%) showed an enrichment trend in R–HB. However, more than half of differentially abundant taxa were enriched in R–HB, which was significantly correlated with organic matter (OM). Bugbase phenotypic predictions indicated a low abundance of the anaerobic phenotype in R–HF and a high abundance of the biofilm-forming phenotype in R–HB.

Novel therapeutic strategies for rare mutations in non-small cell lung cancer

Lung cancer is still the leading cause of cancer-related mortality. Over the past two decades, the management of non-small cell lung cancer (NSCLC) has undergone a significant revolution. Since the first identification of activating mutations in the epidermal growth factor receptor (EGFR) gene in 2004, several genetic aberrations, such as anaplastic lymphoma kinase rearrangements (ALK), neurotrophic tropomyosin receptor kinase (NTRK) and hepatocyte growth factor receptor (MET), have been found. With the development of gene sequencing technology, the development of targeted drugs for rare mutations, such as multikinase inhibitors, has provided new strategies for treating lung cancer patients with rare mutations. Patients who harbor this type of oncologic driver might acquire a greater survival benefit from the use of targeted therapy than from the use of chemotherapy and immunotherapy. To date, more new agents and regimens can achieve satisfactory results in patients with NSCLC. In this review, we focus on recent advances and highlight the new approval of molecular targeted therapy for NSCLC patients with rare oncologic drivers.

4 Nutrition and gut microbial ecology in companion animals

Abstract The gut microbiome, considered an organ harboring trillions of microbes, is a complex and diverse ecosystem through its collective metabolic activities and host interactions that influences host physiology and disease susceptibilities. The advancement from traditional culturable techniques to next generation sequencing technologies allows nutritionists and clinicians to enhance the understanding of microbiomes and their beneficial role in the host-microbe ecosystem. Compared with the research and resource developments in the human microbiome space, the gut microbiome of companion animals is relatively underexplored despite its relevance to animal and pet owner health. More than a decade ago, Hill’s Pet Nutrition developed deep microbiome expertise to improve the overall health and wellbeing for dogs and cats through nutrition. This presentation will focus on two aspects: how Hill’s harnesses the gut microbiome through nutrition to benefit the health of the pet, and the efforts underway to build the “One Health Microbiome Resource (OHMR).” The OHMR resource is a reference database encompassing both human and companion animal metagenomes that will improve the understanding of how microbiomes overlap in shaping the health of pets and pet owners in the context of infectious diseases, immune modulation, and specific genetic elements such as antibiotic resistance genes.

Identifying Metabolic Diseases That Precipitate Neonatal Seizures.

Although a rare cause of neonatal seizures, inborn errors of metabolism (IEMs) remain an essential component of a comprehensive differential diagnosis for poorly controlled neonatal epilepsy. Diagnosing neonatal-onset metabolic conditions proves a difficult task for clinicians; however, routine state newborn screening panels now include many IEMs. Three in particular-pyridoxine-dependent epilepsy, maple syrup urine disease, and Zellweger spectrum disorders-are highly associated with neonatal epilepsy and neurocognitive injury yet are often misdiagnosed. As research surrounding biomarkers for these conditions is emerging and gene sequencing technologies are advancing, clinicians are beginning to better establish early identification strategies for these diseases. In this literature review, the authors aim to present clinicians with an innovative clinical guide highlighting IEMs associated with neonatal-onset seizures, with the goal of promoting quality care and safety.

Germline NGS targeted analysis in adult patients with sporadic adrenocortical carcinoma

BackgroundAdrenocortical carcinoma (ACC) is a rare cancer that arises sporadically or due to hereditary syndromes. Data on germline variants (GVs) in sporadic ACC are limited. Our aim was to characterize GVs of genes potentially related to adrenal diseases in 150 adult patients with sporadic ACC. MethodsThis was a retrospective analysis of stage I-IV ACC patients with sporadic ACC from two reference centers for ACC in Italy. Patients were included in the analysis if they had confirmed diagnosis of ACC, a frozen peripheral blood sample and complete clinical and follow-up data. Next generation sequencing technology was used to analyze the prevalence of GVs in a custom panel of 17 genes belonging to either cancer-predisposition genes or adrenocortical-differentiation genes categories. ResultsWe identified 18 GVs based on their frequency, enrichment and predicted functional characteristics. We found six pathogenic (P) or likely pathogenic (LP) variants in ARMC5, CTNNB1, MSH2, PDE11A and TP53 genes; and twelve variants lacking evidence of pathogenicity. New unique P/LP variants were identified in TP53 (p.G105D) and, for the first time, in ARMC5 (p.P731R). The presence of P/LP GVs was associated with reduced survival outcomes and had a significant and independent impact on both progression-free survival and overall survival. ConclusionsGVs were present in 6.7 % of patients with sporadic ACC, and we identified novel variants of ARMC5 and TP53. These findings may improve understanding of ACC pathogenesis and enable genetic counseling of patients and their families.

Thoracic spine infection caused by Pseudomonas fluorescens: A case report and review of literature

BACKGROUND The clinical incidence of spinal infection is gradually increasing, and its onset is insidious, easily leading to missed diagnosis and misdiagnosis, which may lead to serious complications such as nervous system dysfunction, spinal instability and/or deformity, and cause a huge burden on society and families. Early identification of the causative agent and precision medicine will greatly reduce the suffering of patients. At present, the main pathogenic bacteria that cause spinal infection are Staphylococcus aureus , Streptococcus, Pneumococcus, Escherichia coli , and Klebsiella. There are no reports of spinal infection caused by Pseudomonas fluorescens . CASE SUMMARY We report a 32-year-old female patient with spinal infection. She presented with flank pain, initially thought to be bone metastases or bone tuberculosis, and had a family background of tumors. Her clinical features and changes in imaging and laboratory tests led to the suspicion of thoracic spine infection. Histopathology of the lesion showed inflammation, tissue culture of the lesion was negative several times, and the possible pathogen - Pseudomonas fluorescens was found after gene sequencing of the lesion. The patient recovered completely after a full course of antibiotic treatment. CONCLUSION This report increases the range of pathogens involved in spinal infections, highlights the unique advantages of gene sequencing technology in difficult-to-diagnose diseases, and validates conservative treatment with a full course of antibiotics for spinal infections without complications.