scholarly journals Metagenomic next-generation sequencing diagnosis of peripheral pulmonary infectious lesions through virtual navigation, radial EBUS, ultrathin bronchoscopy, and ROSE

2019 ◽  
Vol 47 (10) ◽  
pp. 4878-4885 ◽  
Author(s):  
Nana Liu ◽  
Jianying Kan ◽  
Wenbin Cao ◽  
Jie Cao ◽  
Erlie Jiang ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Lung Infection ◽  
Endobronchial Ultrasound ◽  
Pathogenic Microorganisms ◽  
Next Generation ◽  
Specificity And Sensitivity ◽  
Peripheral Lung ◽  
Site Evaluation ◽  
Generation Sequencing ◽  
Higher Sensitivity

Objective To evaluate the efficacy of combined rapid on-site evaluation of cytology (ROSE), ultrathin bronchoscopy, virtual bronchoscopic navigation, radial endobronchial ultrasound (EBUS), and metagenomic next-generation sequencing (mNGS) for diagnosis of peripheral pulmonary infectious lesions. Methods Specimens from patients with peripheral lung infection were obtained by transbronchial lung biopsy (TBLB) and bronchoalveolar lavage (BAL), and mNGS was used to detect pathogenic microorganisms. The sensitivity and specificity of mNGS were compared between TBLB tissue and BAL fluid. Results The most common pathogens of pulmonary infectious lesions in this study were Pseudomonas aeruginosa, Klebsiella pneumoniae, and Acinetobacter baumannii. The specificity of mNGS was higher in TBLB tissue than in BAL fluid, but mNGS of BAL fluid had higher sensitivity. Conclusions The combination of ROSE, ultrathin bronchoscopy, virtual bronchoscopic navigation, radial EBUS, and mNGS technology yielded high efficacy for the diagnosis of peripheral pulmonary infectious lesions. TBLB and BAL specimens have respective advantages in specificity and sensitivity for mNGS analysis.

scholarly journals The Value of Combined Radial Endobronchial Ultrasound-Guided Transbronchial Lung Biopsy and Metagenomic Next-Generation Sequencing for Peripheral Pulmonary Infectious Lesions

2020 ◽  
Vol 2020 ◽  
pp. 1-9
Author(s):  
Guangsheng Li ◽  
Jie Huang ◽  
Yuechuan Li ◽  
Jing Feng
Keyword(s):  
Next Generation Sequencing ◽  
Confidence Interval ◽  
Odds Ratio ◽  
Lung Biopsy ◽  
Pathogen Detection ◽  
Lung Infection ◽  
Endobronchial Ultrasound ◽  
Image Characteristics ◽  
Peripheral Lung ◽  
Generation Sequencing

Background. Metagenomic next-generation sequencing (mNGS) is a new technology that allows for unbiased detection of pathogens. However, there are few reports on mNGS of lung biopsy tissues for pulmonary infection diagnosis. In addition, radial endobronchial ultrasound (R-EBUS) is widely used to detect peripheral pulmonary lesions (PPLs), but it is rarely used in the diagnosis of peripheral lung infection. Objective. The present study aims to evaluate the combined application of R-EBUS-guided transbronchial lung biopsy (TBLB) and mNGS for the diagnosis of peripheral pulmonary infectious lesions. Methods. From July 2018 to April 2019, 121 patients from Tianjin Medical University General Hospital diagnosed with PPLs and lung infection were enrolled in this prospective randomized study . Once the lesion was located, either TBLB or R-EBUS-guided-TBLB was performed in randomly selected patients, and mNGS was applied for pathogen detection in lung biopsy tissues. The results of mNGS were compared between the TBLB group and R-EBUS-guided TBLB group. In addition, the clinical characteristics and EBUS images from 61 patients receiving bronchoscopy for peripheral lung infectious detection were analyzed and compared with the results of mNGS. Results. The positivity rate of mNGS in R-EBUS-guided TBLB was (78.7%, 48/61) that was significantly higher than (60.0%, 36/60) in the TBLB group. Difference in the position of R-EBUS probe and image characteristics of peripheral lung infectious lesions affected the positivity rate of mNGS. Tissue collected by R-EBUS within the lesion produced higher positivity rate than samples collected adjacent to the lesion (P=0.030, odds ratio 17.742; 95% confidence interval, from 1.325 to 237.645). Anechoic areas and luminant areas of ultrasonic image characteristics were correlated with lower positivity rate of mNGS (respectively, P=0.019, odds ratio 17.878; 95% confidence interval, from 1.595 to 200.399; P=0.042, odds ratio 16.745; 95% confidence interval, from 1.106 to 253.479). Conclusions. R-EBUS-guided TBLB is a safe and effective technique in the diagnosis of peripheral lung infectious lesions. R-EBUS significantly facilitates the accurate insertion of bronchoscope into the lesions, which improves positivity rate of mNGS analysis in pathogen detection. The R-EBUS probe position within lesion produced a higher positivity rate of mNGS analysis. Nevertheless, the presence of anechoic and luminant areas on ultrasonic image was correlated with poor mNGS positivity rate.

scholarly journals Suitability of Bronchoscopic Biopsy Tissue Samples for Next-Generation Sequencing

Diagnostics ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 391
Author(s):  
Shuji Murakami ◽  
Tomoyuki Yokose ◽  
Daiji Nemoto ◽  
Masaki Suzuki ◽  
Ryou Usui ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Success Rate ◽  
Rna Sequencing ◽  
Surface Area ◽  
Endobronchial Ultrasound ◽  
Next Generation ◽  
Endobronchial Ultrasonography ◽  
Tissue Surface ◽  
The Impact ◽  
Generation Sequencing

A sufficiently large tissue sample is required to perform next-generation sequencing (NGS) with a high success rate, but the majority of patients with advanced non-small-cell lung cancer (NSCLC) are diagnosed with small biopsy specimens. Biopsy samples were collected from 184 patients with bronchoscopically diagnosed NSCLC. The tissue surface area, tumor cell count, and tumor content rate of each biopsy sample were evaluated. The impact of the cut-off criteria for the tissue surface area (≥1 mm2) and tumor content rate (≥30%) on the success rate of the Oncomine Dx Target Test (ODxTT) was evaluated. The mean tissue surface area of the transbronchial biopsies was 1.23 ± 0.85 mm2 when small endobronchial ultrasonography with a guide sheath (EBUS-GS) was used, 2.16 ± 1.49 mm2 with large EBUS-GS, and 1.81 ± 0.75 mm2 with endobronchial biopsy (EBB). The proportion of samples with a tissue surface area of ≥1 mm2 was 48.8% for small EBUS-GS, 79.2% for large EBUS-GS, and 78.6% for EBB. Sixty-nine patients underwent ODxTT. The success rate of DNA sequencing was 84.1% and that of RNA sequencing was 92.7% over all patients. The success rate of DNA (RNA) sequencing was 57.1% (71.4%) for small EBUS-GS (n = 14), 93.4% (96.9%) for large EBUS-GS (n = 32), 62.5% (100%) for EBB (n = 8), and 100% (100%) for endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) (n = 15). Regardless of the device used, a tissue surface area of ≥ 1 mm2 is adequate for samples to be tested with NGS.

scholarly journals Suitability of transbronchial brushing cytology specimens for next‐generation sequencing in peripheral lung cancer

2020 ◽  
Vol 112 (1) ◽  
pp. 380-387
Author(s):  
Naoki Furuya ◽  
Shingo Matsumoto ◽  
Kazutaka Kakinuma ◽  
Kei Morikawa ◽  
Takeo Inoue ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Next Generation Sequencing ◽  
Next Generation ◽  
Peripheral Lung Cancer ◽  
Brushing Cytology ◽  
Peripheral Lung ◽  
Generation Sequencing

scholarly journals Detection and identification of Acanthamoeba and other non-viral causes of infectious keratitis in corneal scrapings by real-time PCR and next-generation sequencing-based 16S-18S gene analysis

2020 ◽  
Author(s):  
Dennis Back Holmgaard ◽  
Celine Barnadas ◽  
Seyed Hossein Mirbarati ◽  
Lee O’Brien Andersen ◽  
Henrik Vedel Nielsen ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Real Time ◽  
Real Time Pcr ◽  
Clinical Relevance ◽  
Bacterial Species ◽  
Screening Method ◽  
Infectious Keratitis ◽  
Next Generation ◽  
Specificity And Sensitivity ◽  
Generation Sequencing

Acanthamoeba is a free-living amoeba of extensive genetic diversity. It may cause infectious keratitis (IK), which can also be caused by bacteria, fungi, and viruses. High diagnostic sensitivity is essential to establish an early diagnosis of Acanthamoeba-associated keratitis. Here, we investigated the applicability of next-generation sequencing (NGS)-based ribosomal gene detection and differentiation (16S-18S) compared with specific real-time PCR for detection of Acanthamoeba. Two hundred DNAs extracted from corneal scrapings and screened by Acanthamoeba-specific real-time PCR were analyzed using an in-house 16S-18S NGS assay. Of these, 24 were positive using specific real-time PCR, 21 of which were positive using the NGS assay. Compared with real-time PCR; the specificity and sensitivity of the NGS assay were 100% and 88%, respectively. Genotypes identified by the NGS assay included T4 (n = 19) and T6 (n = 2). Fungal and bacterial species of potential clinical relevance were identified in 31 of the samples negative for Acanthamoeba, exemplified by Pseudomonas aeruginosa (n = 11), Moraxella spp. (n = 6), Staphylococcus aureus (n = 2), Fusarium spp. (n = 4), and Candida albicans (n = 1). Conclusively, the 16S-18S assay was slightly less sensitive than real-time PCR in detecting Acanthamoeba-specific DNA in corneal scrapings. Robust information on genotype was provided by the NGS assay, and other pathogens of potential clinical relevance were identified in 16% of the samples negative for Acanthamoeba. NGS-based detection of ribosomal genes in corneal scrapings could be an efficient screening method for detecting non-viral causes of IK, including Acanthamoeba.

scholarly journals Gestational Psittacosis With Secondary Hemophagocytic Syndrome: A Case Report and Literature Review

2021 ◽  
Vol 8 ◽  
Author(s):  
Li Sun ◽  
Pulin Li ◽  
Bo Pang ◽  
Peipei Wu ◽  
Ran Wang
Keyword(s):  
Bone Marrow ◽  
Next Generation Sequencing ◽  
Hemophagocytic Syndrome ◽  
Nk Cell ◽  
Cell Activity ◽  
Lung Infection ◽  
Next Generation ◽  
Nk Cell Activity ◽  
Generation Sequencing

Gestational psittacosis and hemophagocytic syndrome (HPS) are rare clinical diseases. In this article, a case of gestational psittacosis concomitant with secondary HPS was reported. An analysis was performed on the clinical characteristics, signs, laboratory findings, progression, diagnosis, and treatment of a patient with gestational psittacosis concomitant with secondary HPS. Besides, the literature with respect to this disease was reviewed. This patient was definitively diagnosed through metagenomic next-generation sequencing techniques, bone marrow puncture and smear examination, and the determination of sCD25 level and natural killer (NK) cell activity. Anti-infectives such as doxycycline and etoposide combined with hormone chemotherapy achieved significant improvement in cough and expectoration, a return to normal temperature, and a significant improvement in oxygenation index. In addition, chest computed tomography revealed obvious absorption of lung lesions and a return of NK cell activity and sCD25 levels to normal ranges. Chlamydia psittaci pneumonia requires a clear determination of etiology, while HPS requires bone marrow puncture and smear examination, together with the determination of sCD25 level and NK cell activity in the blood. The findings of this study suggest that metagenomic next-generation sequencing is an effective instrument in clearly identifying pathogens that cause lung infection. Clinicians should consider atypical pathogens of lung infection in patients with poor response to empirical anti-infectives, and strive to design an effective treatment strategy as per an accurate diagnosis based on the etiology. As for patients suffering from long-term high fever and poor temperature control after broad-spectrum antibiotic treatment, non-infectious fever should be taken into account. A rapid and clear diagnosis would significantly improve patient prognosis.

Feasibility of endobronchial ultrasound fine-needle aspiration for massively parallel next-generation sequencing in cancer patients.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. 8535-8535
Author(s):  
Simon R. Turner ◽  
Darren Buonocore ◽  
Natasha Rekhtman ◽  
Snjezana Dogan ◽  
Patrice Desmeules ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Fine Needle Aspiration ◽  
Needle Aspiration ◽  
Endobronchial Ultrasound ◽  
Precision Oncology ◽  
Next Generation ◽  
Cell Free Dna ◽  
Fine Needle ◽  
Free Dna ◽  
Generation Sequencing

8535 Background: Next generation sequencing (NGS) is an important emerging tool in precision oncology, allowing identification of a growing number of clinically validated and investigational therapeutic molecular targets. A potential limitation is that some NGS assays require more DNA input than more limited molecular assays. Endobronchial ultrasound fine-needle aspiration (EBUS-FNA) is a minimally invasive procedure for sampling mediastinal and pulmonary lesions, but it is unknown if it provides adequate material for NGS. Methods: An IRB approved, retrospective review was performed of patients undergoing EBUS-FNA by thoracic surgeons at our institution 3/1/14 - 9/28/16. NGS was performed using an assay developed at our institution that detects mutations in up to 410 genes (MSK-IMPACT). Samples diagnostic for malignancy and with MSK-IMPACT requested were identified. Pathology and clinical data were drawn from the medical record and MSK-IMPACT results were examined. Results: 784 EBUS-FNA were done in the study period. MSK-IMPACT was requested on 115 positive samples. MSK-IMPACT was successful in 99 samples (86.1%), identifying an average of 12.7 mutations at a mean coverage depth of 806X. In 17 (17.2%) samples, tumor content was suboptimal ( < 20% of nucleated cells), with fewer identified mutations than in cases with higher tumor content (6.8 vs 13.9, p = 0.01). NGS was performed on paraffin-embedded cell blocks in 93 cases (93.9%), and in 6 cases DNA extraction was performed from residual cytological material isolated from supernatant including cell-free DNA. Failures were attributable to low cell content (7), high contamination by benign cells (4) or both (1) and processing issues (4). No difference in surgical or radiologic parameters were identified for failed or suboptimal samples. Conclusions: In our practice, EBUS-FNA has a high rate of success for obtaining adequate tissue for NGS. Ability to utilize cell-free DNA for molecular studies – a new process in our lab – allows increased success of molecular testing in scant samples. Further studies may identify factors contributing to NGS failure and to improving success for samples with minimal cellularity.

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