Objective To analyze the differences in microbial communities in bronchoalveolar lavage fluid (BALF) from patients with simple pneumonia versus those with chronic obstructive pulmonary disease (COPD) combined with lower respiratory tract infection using metagenomic next-generation sequencing (mNGS). Methods Patients hospitalized for pulmonary infections at the First Affiliated Hospital of Xinjiang Medical University between December 2021 and March 2023 were included. Based on the presence of COPD, the patients were divided into two groups: those with simple pneumonia and those with COPD combined with lower respiratory tract infection. mNGS was employed to detect microbes in BALF, and the microbial community distribution characteristics of the two groups were analyzed. Results A total of 97 patients were included, of whom 80 (81.82%) had positive microbial detection results. The smoking index in COPD group with lower respiratory tract infection was significantly higher than that in the group with simple pneumonia (t= −3.62, P=0.001). Differences in microbial community distributions were observed between the groups. At the genus level, 19 species of microorganisms were detected in the simple pneumoniapulmonary infection group, including 8 bacteria (42.11%), 2 fungi (10.53%), 3 viruses (15.79%), and 6 other types of microorganisms (31.58%). In contrast, 22 types of microbes were detected in COPD group with lower respiratory tract infection, including 10 bacteria (47.62%), 3 fungi (14.29%), 4 viruses (19.05%), and 4 other types of microorganisms (19.05%). Differences were also noted in reads per million (RPM) values; bacterial RPM values at the genus level were significantly higher in the COPD group during non-severe pneumonia compared to the simple pneumonia group (Z=–2.706, P=0.007). In the patients with severe pneumonia, RPM values at the genus and species levels were significantly higher than those in non-severe pneumonia (Z=−2.202, P=0.028; Z=−2.141, P=0.032). In COPD combined with severe pneumonia, bacterial RPM values were significantly higher at the species level compared to non-severe pneumonia (Z=−2.367, P=0.017). ConclusionsThere are differences in the distribution of microbial communities at the genus and species levels in BALF from patients with COPD combined with lower respiratory tract infection compared to those with simple pulmonary pneumonia. Bacteria are the predominant microbial type in both groups, but the dominant bacterial species differ between them. Simple pneumonia are primarily associated with bacterial, viral, and other types of microbial infections, while COPD combined with lower respiratory tract infection is predominantly associated with fungal and bacterial infections. RPM values may serve as an indicator of the severity of pneumonia.
In recent years, due to the extensive usage of immunosuppressant and the rise of patients with cancers and organ transplantation, the incidence rate of invasive fungal infection, especially invasive pulmonary fungal infection, has increased. Besides the clinical manifestations, medical history and imaging, the diagnosis of pulmonary mycosis mainly depends on pathogen detection methods in clinical microbiology laboratory. However, due to the difficulty in fungi culturing and the low sensitivity of smear microscopy, better molecular biology methods are needed. To date, the emergence of metagenomic next-generation sequencing (mNGS) has improved the identification rate of pulmonary fungal infections. mNGS is significantly superior to traditional detection methods in rapid, accurate, and comprehensive determination of fungi from various clinical specimens, especially atypical fungi. However, some problems in mNGS method have to be addressed including sample collection, report interpretation, and its combination with traditional microbiology methods. With the in-depth discussion and solution of the above problems, mNGS will be indispensable to the etiological diagnosis of pulmonary invasive fungal infection.
ObjectiveTo study the application of non-real-time ultrasound bronchoscopy combined with Metagenomic Next-Generation Sequencing (mNGS) for diagnosis in focal pulmonary infectious diseases. MethodsProspective inclusion of patients with focal pulmonary infection were randomly divided into two groups, the experimental group used non-real-time ultrasound bronchoscopy positioning to collect bronchial alveolar lavage fluid (BALF), while the control group used chest CT position. BALF was subjected to mNGS and traditional microbial detection including traditional culture, the fungal GM test and Xpert (MTB/RIF). ResultThe positive rate of traditional culture (39.58% vs. 16.67%, P=0.013) and mNGS (89.58% vs. 72.92%, P=0.036) in experimental group was higher. The positive rate of Xpert MTB/RIF (4.17% vs. 2.08%, P=1) and fungal GM test (6.25% vs. 4.17%, P=0.765) was similar. The positive rate of bacteria and fungi detected by mNGS was higher than traditional culture (61.46% vs. 28.13%, P<0.001). Mycobacterium tuberculosis was similar to Xpert MTB/RIF (8.33% vs. 3.13%, P=0.21). Aspergillus was similar to GM test (7.29% vs. 5.21%, P=0.77). The total positive rate of traditional microbial methods was 36.46%, but 81.25% in mNGS (P<0.001). mNGS showed that 35 cases were positive and 13 kinds of pathogens were detected in control group, but 43 patients and 17 kinds of pathogens were detected in experimental group. The average hospitalization time [(12.92±3.54) days vs. (16.35±7.49) days] and the cost [CNY (12209.17±3956.17) vs. CNY (19044.10±17350.85)] of experimental group was less (P<0.001). ConclusionsNon-real-time ultrasound bronchoscopy combined with mNGS can improve the diagnostic rate of focal pulmonary infectious diseases which is worthy of popularization and application in clinical practice.
In recent years, with the wide application of metagenomics next-generation sequencing, more and more rare pathogens have been detected in our clinical work, including non-tuberculous Mycobacterium, Corynebacterium, Fusarium, Cryptococcus pneumoniae, human herpes virus, torque teno virus, parvovirus, Tropheryma whipplei, Bartonella, Chlamydia psittaci, etc. It is difficult to determine whether these rare pathogens are clinically significant and need treatment. This article puts forward some suggestions and discussions on the diagnosis and treatment of pulmonary infections with some rare pathogens.
Objective To explore the application value of metagenomic next-generation sequencing (mNGS) based on human sequencing in the clinical early diagnosis of lung cancer. Methods Four patients hospitalized with suspected lung infection were retrospectively analyzed, and the test results of bronchoalveolar lavage fluid (BALF) on mNGS of tumor metagenome, the routine clinical test results, and their clinical diagnosis and treatment information in between August 26, 2021, and December 18, 2021. Results Patient 1 was preliminarily diagnosed with lung cancer by referring to chest computed tomography (CT) imaging. Chest radiograph or CT in the other three patients showed bilateral lung CT and lamellar hyperintensities (patient 2), bilateral lung mass-like and lamellar hyperintensities (patient 3), and lung masses (patient 4), respectively. BALF samples from all 4 patients were detected with mNGS based on human tumor sequences, indicating tumor. In addition, the result in patient 3 also indicated white pseudofilamentous yeast infection consistent with clinical culture, and the result in patient 4 also showed infection of rhinovirus type A. Conclusion The second generation genome sequencing technology based on human sequence can not only assist clinical diagnosis of infection, but also provide detection datUM support for tumor early warning.
Objective To analyze the difference of sputum flora between acute exacerbation and stable chronic obstructive pulmonary disease (COPD) patients basing on metagenomic next generation sequencing (mNGS), and its relationship with clinical indicators. The role of sputum flora of COPD patients in unexplained deterioration was explored, so as to find a targeted treatment plan. Methods From December 2021 to June 2022, 54 COPD patients who had a history of smoking were recruited, including 25 patients in stable COPD (SCOPD group) and 29 patients in acute exacerbation (AECOPD group). The sputum was collected and sequenced by mNGS, and the difference of sputum flora between the two groups was compared. Results Compared with SCOPD group, the evenness of sputum flora (Shannon index) in AECOPD group decreased significantly (P=0.019, Mann-Whitney U test). At the phylum level, the relative abundance of Fusobacteria in AECOPD group was significantly lower than that in SCOPD group (Z=–2.669, P=0.008). At genus level, compared with SCOPD group, the relative abundance of Fusobacterium and Haemophilus in AECOPD group decreased significantly (Z=–3.062, P=0.002; Z=–2.143, P=0.032), and the relative abundance of Granulicatella increased significantly (Z=–2.186, P=0.029). At species level, the relative abundance of sputum Haemophilus parainfluenzae, Moraxella catarrhalis and Haemophilus influenzae in AECOPD group was significantly lower than that in SCOPD group (Z=–2.230, P=0.026; Z=–2.125, P=0.034; Z=–2.099, P=0.036). At the time of acute exacerbation of COPD, the relative abundance of Gemella in sputum was positively correlated with forced expiratory volume in first second/forced vital capacity (FEV1/FVC) and body mass index (r=0.476, P=0.009; r=0.427, P=0.021), which was negatively correlated with nutrition risk screening 2002 (r=–0.570, P=0.001). The relative abundance of Neisseria and Neisseria subflava was negatively correlated with GOLD grade (r=–0.428, P=0.020; r=–0.455, P=0.013). The relative abundance of Rothia aeria was posotively correlated with C-reactive peotein (r=0.388, P=0.038). Conclusions There are significant differences of sputum flora in phylum, genus and species level between stable and acute exacerbation COPD patients. The evenness of sputum flora in COPD patients in acute exacerbation is significantly lower than that in patients in stable stage. Fusobacteria, Fusobacterium, Gemella and Nesseria (Neisseria subflava) may play a beneficial role in COPD, while Rothia aeria may be associated with COPD exacerbation.
ObjectiveTo explore the application and clinical value of metagenomic next-generation sequencing (mNGS) combined with Omadacycline in the treatment of Refractory Mycoplasma pneumoniae pneumonia (RMPP).MethodsThe clinical data, relevant laboratory results, diagnosis and treatment process, and imaging outcomes of four patients diagnosed with Mycoplasma pneumoniae pneumonia through mNGS were analyzed. ResultsThe clinical symptoms at onset in all four patients were consistent with Mycoplasma pneumoniae pneumonia. After conventional treatment with macrolides, tetracyclines, or quinolone antibiotics, the symptoms showed no significant improvement, and there was a trend of radiological worsening. Following the confirmation of Mycoplasma pneumoniae infection through mNGS of bronchoalveolar lavage fluid, and due to various reasons preventing the use of the aforementioned drugs, omadacycline was ultimately chosen for treatment. Radiological improvements were observed in all cases, leading to a good prognosis and discharge. ConclusionsFor pneumonia cases where the infectious pathogen cannot be identified and conventional treatment has failed, mNGS can be utilized for early and accurate diagnosis. In cases of RMPP, Omadacycline can be employed as an alternative treatment to prevent delays in care and reduce the risk of complications.
ObjectiveTo improve the understanding of psittacosis, the clinical data of 8 cases are reviewed. The application of pathogen metagenomics next-generation sequencing (mNGS) in the diagnosis of nocardiosis is also investigated.MethodsThe clinical data of eight patients with psittacosis diagnosed by mNGS in Nanjing Drum Tower Hospital from January 2018 to May 2020 were reviewed. The clinical characteristics, laboratory examination characteristics and imaging changes were analyzed, and the treatment outcome was followed-up.ResultsAmong the eight cases, there were six males and two females, aged 43～83 years old, with an average age of 64±12 years old. Six of them had a clear history of poultry exposure. The major clinical manifestations were fever, cough, dyspnea, etc. Chest high-resolution computed tomography (HRCT) may have solid shadow, ground glass like shadow. Chlamydia psittaci was detected by mNGS in eight patients’ bronchoalveolar lavage fluid. Minocycline or moxifloxacin were administrated, six patients were discharged after their condition improved, and two patients died.ConclusionsThe incidence of psittacosis is low, and its clinical manifestations lack specificity. In the course of the disease, there may be different degrees of fever, cough, sputum, dyspnea and other symptoms. The lungs can be heard with wet rales, chest HRCT can be seen ground glass shadow, consolidation shadow, accompanied by air bronchogram. Chlamydia psittaci can be detected in alveolar lavage fluid by mNGS. The patients need to be treated for a long time, lasting at least 10 to 14 days. Tetracycline drugs should be the first choice, and can be combined with other antibiotics with activity against gram-positive and gram-negative bacteria in critical patients.