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Objective:To investigate the diagnostic value of nucleic acid matrix-assisted laser desorption ionization-time of flight mass spectrometry(MALDI-TOF MS)in sputum smear-negative patients with nontuberculous Mycobacterial(NTM)pulmonary disease.Methods:Clinical data of 123 patients suspected of NTM pulmonary disease admitted in Public Health Clinical Center Affiliated to Shandong University between July 2022 and November 2023 were retrospectively analyzed. Bronchoalveolar lavage fluid(BALF)specimens were collected for MALDI-TOF MS assay and MGIT 960 culture. The diagnostic efficacy of MALDI-TOF MS for NTM pulmonary disease in patients with negative sputum smears for acid-fast bacilli was evaluated with receiver operating characteristic(ROC)curve. Statistical analysis was performed using SPSS 26.0 software and MedCalc statistical software.Results:Diagnosis of NTM pulmonary disease was finally confirmed in 66 out of the 123 suspected patients. It took 8 to 24 h for MALDI-TOF MS to identify NTM species and resistance. By MALDI-TOF MS,72 NTM strains were identified,with the Mycobacterium avium complex being the most prevalent(34 strains,47.22%),followed by the Mycobacterium abscessus complex(13 strains,18.06%);resistance to macrolides was detected in 6 cases,while no resistance to aminoglycosides was found. It took 9 to 45 days for BALF MGIT 960 culture to identify NTM,and took 7 to 15 days for NTM typing and drug sensitivity testing. By BALF MGIT 960 culture,28 NTM strains were identified;and 1 case was found to be resistant to macrolides. Using confirmed diagnosis as the gold standard,MALDI-TOF MS demonstrated higher sensitivity,negative predictive value,and agreement rate compared to MGIT 960 culture(84.85% vs. 42.42%,81.13% vs. 56.32%,80.49% vs. 62.60%, χ2=25.667,8.998,9.664, P<0.05 or <0.01). The area under ROC curve(AUC)for MALDI-TOF MS was significantly higher than that of MGIT 960 culture(0.801 vs. 0.642, Z=3.300, P=0.001). Conclusion:Compared to MGIT 960 culture,MALDI-TOF MS exhibits superior diagnostic efficiency in detecting NTM pulmonary disease in patients with acid-fast bacilli smear-negative sputum,with advantage of rapidly identifying NTM species and resistance.
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Background It is a research hotspot to study the changes of metabolites and metabolic pathways in the process of coal worker's pneumoconiosis (CWP) by metabonomics and to explore its pathogenesis. Objective To study the change of metabolites in bronchoalveolar lavage fluid (BALF) of patients with CWP and explore the metabolic regulation mechanism of the disease. Methods Patients with CWP who met the national diagnostic criteria according to Diagnosis of occupational pneumoconiosis (GBZ 70-2015) and underwent massive whole lung lavage were selected as the case group, and patients with tracheostenosis who underwent bronchoscopy were selected as the control group. BALF samples were collected from the cases and the controls. After filtering out large particles and mucus, the supernatant was stored in a −80 ℃ refrigerator. The samples were detected and analyzed by liquid chromatography-mass spectrometry after adding extraction solution, cold bath ultrasonication, and high-speed centrifugation, and the metabolic profiles and related data of CWP patients were obtained. The differential metabolites related to the occurrence and development of CWP were screened by multiple statistical analysis; furthermore, we searched the Kyoto Encyclopedia of Genes and Genomes (KEGG) database for potential metabolic pathways involved in the progression. Results There was no significant difference in the general conditions of the subjects, such as weight, height, age, and length of service among the stage I group, the stage II group, the stage III group, and the control group (P˃0.05). When comparing the CWP stage I group with the control group, 48 differential metabolites were screened out, among which 14 were up-regulated and 34 were down-regulated. A total of 66 differential metabolites were screened out between the patients with CWP stage II and the controls, 14 up-regulated and 52 down-regulated differential metabolites. Compared with the control group, 63 differential metabolites were screened out in the patients with CWP stage III, including 11 up-regulated and 52 down-regulated differential metabolites. There were 36 differential metabolites that may be related to the occurrence of CWP, among which 11 differential metabolites were up-regulated, and 25 were down-regulated. Four significant differential metabolic pathways were identified through KEGG database query: linoleic acid metabolic pathway, alanine metabolic pathway, sphingolipid metabolic pathway, and glycerophospholipid metabolic pathway. Conclusion The metabolomic study of BALF show that there are 36 different metabolites in the occurrence and development of CWP, mainly associating with linoleic acid metabolism, alanine metabolism, sphingolipid metabolism, and glycerophospholipid metabolism pathways.
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Objective:To establish a performance validation method for mNGS applied in BALF samples.Method:Hela cells were used as a representative of host cells, and simulated BALF samples were prepared by adding different concentrations of Hela cells, seven species of isolated pathogens (including Streptococcus pneumonia, Hemophilus influenza, Klebsiella pneumonia, Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, and Adenovirus), and interfering substances to sterile normal saline. Clinical BALF samples were collected simultaneously, and the results of mNGS were evaluated using traditional detection methods as a reference. The limit of detection (LOD), precision, anti-interference ability, stability, and accuracy of mNGS were determined. Results:In the simulated samples, the LOD of Streptococcus pneumoniae, Haemophilus influenzae, Klebsiella pneumoniae, Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, and Adenovirus were 150, 262, 102, 67, 96, 83 CFU/ml, and 439 copies/ml, respectively. The repeatability of the detection results for all pathogens of simulated positive BALF samples was 100%. The anti-interference test showed that the higher the concentration of human DNA, the fewer pathogen sequences detected by mNGS. Escherichia coli and Shigella sonnei were used to evaluate the ability of mNGS to distinguish closely related species. The results showed that the system could stably distinguish Escherichia coli and Shigella sonnei when the concentration of Shigella sonnei was 4, 000 CFU/ml. The stability test results showed that there was no significant change in the number of pathogen sequences detected whether after 1 to 3 freeze-thaw cycles or storage at 4 ℃, -20 ℃, or -80 ℃ for 36 h. Compared with traditional detection methods, the accuracy of 17 clinical samples was 82.4%(14/17). Continuous evaluation of clinical BALF samples simultaneously tested by mNGS and traditional methods at Tongji Hospital from October 25, 2021, to September 14, 2022, showed that the accuracy of mNGS compared to bacterial culture, fungal culture, mycobacterial culture, Mycobacterium tuberculosis culture, and conventional PCR techniques was 67.5%(472/699), 81.5%(570/699), 92.3%(335/363), 96.4%(350/363), and 86.8%(132/152), respectively. Compared with conventional PCR techniques, the accuracy of mNGS for detecting Pneumocystis jirovecii, Adenovirus, and Mycoplasma pneumoniae was 89.4%(84/94), 93.3%(56/60), and 87.1%(61/70), respectively. Conclusion:By preparing simulated BALF samples and using traditional detection methods as a reference, the performance characteristics of mNGS in detecting BALF samples can be preliminarily evaluated.
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@#Abstract:Objective To investigate the morphological features of the Pneumocystis jirovecii, in order to facilitate early detection and rapid diagnosis of this rare pathogen from a morphology point of view by laboratory technicians. By analyzing the laboratory features and application value of different pathogen detection methods in the diagnosis of Pneumocystis jirovecii pneumonia, we aim to provide the most reliable diagnostic basis for rapid diagnosis of Pneumocystis jirovecii pneumonia.Methods A retrospective analysis was conducted on the test results of bronchoalveolar lavage fluid samples from a comprehensive hospital in Zhangqiu District, Jinan City, Shandong Province, and a hospital in Changde City from April 2022 to October 2022. Five confirmed cases of Pneumocystis jirovecii pneumonia were detected. Its clinical manifestations, laboratory results, and morphological characteristics of pathogens under different stains were analyzed to discuss the advantages and disadvantages of different detection methods. Results Cytological examination of bronchoalveolar lavage fluid found the trophozoites and cysts of Pneumocystis jirovecii by Wright's-Giemsa staining in 4 cases (80%), and the cysts of Pneumocystis jirovecii by Silver hexamine staining in 4 cases (80%), while the metagenomic next-generation sequencing confirmed all the 5 positive results. All 5 patients had different degrees of reduction in the absolute count of peripheral blood lymphocytes, and the serum lactic dehydrogenase and (1-3)-β-D-Glucan were increased. Among the 5 patients in this study, 4 were treated with sulfamethoxazole combined with caspofungin, and 1 was treated with sulfamethoxazole. Three patients were cured and discharged from hospital after treatment, but two died. Conclusions The method of Wright's-Giemsa staining for the cytological examination of bronchoalveolar lavage fluid to find Pneumocystis jirovecii has the unique and irreplaceable advantages as silver staining. Metagenomic next-generation sequencing can further increase the positive detection rate of Pneumocystis jirovecii. The combination of cytological examination of bronchoalveolar lavage fluid with metagenomic nextgeneration sequencing is a powerful diagnostic method for rapid diagnosis of Pneumocystis jirovecii pneumonia, which can diagnose accurately and reduce missed diagnosis.
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The Omicron family of SARS-CoV-2 variants are currently driving the COVID-19 pandemic. Here we analyzed the clinical laboratory test results of 9911 Omicron BA.2.2 sublineages-infected symptomatic patients without earlier infection histories during a SARS-CoV-2 outbreak in Shanghai in spring 2022. Compared to an earlier patient cohort infected by SARS-CoV-2 prototype strains in 2020, BA.2.2 infection led to distinct fluctuations of pathophysiological markers in the peripheral blood. In particular, severe/critical cases of COVID-19 post BA.2.2 infection were associated with less pro-inflammatory macrophage activation and stronger interferon alpha response in the bronchoalveolar microenvironment. Importantly, the abnormal biomarkers were significantly subdued in individuals who had been immunized by 2 or 3 doses of SARS-CoV-2 prototype-inactivated vaccines, supporting the estimation of an overall 96.02% of protection rate against severe/critical disease in the 4854 cases in our BA.2.2 patient cohort with traceable vaccination records. Furthermore, even though age was a critical risk factor of the severity of COVID-19 post BA.2.2 infection, vaccination-elicited protection against severe/critical COVID-19 reached 90.15% in patients aged ≽ 60 years old. Together, our study delineates the pathophysiological features of Omicron BA.2.2 sublineages and demonstrates significant protection conferred by prior prototype-based inactivated vaccines.
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@#Abstract: Objective To investigate the diagnostic value of joint detection of Mycobacterium tuberculosis rifampicin resistance gene (Xpert MTB/RIF), Mycobacterium tuberculosis ribonucleic acid (TB-RNA) and Mycobacterium tuberculosis deoxyribonucleic acid (TB-DNA) in bronchoalveolar lavage fluid for smear-negative pulmonary tuberculosis. Methods A total of 806 patients with suspected smear-negative pulmonary tuberculosis admitted to our hospital from May 2020 to July 2022 were selected, 506 patients diagnosed as bacterial negative pulmonary tuberculosis by clinical, X-ray and sputum samples were classified as bacterial negative pulmonary tuberculosis group, and the other 300 patients with non-tuberculous pulmonary disease were classified as non-tuberculous pulmonary disease group. XpertMTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid of all patients were detected. With clinical, X-ray and sputum specimen examination of mycobacterium tuberculosis as the gold standard, the diagnostic efficacy of alveolar lavage solution Xpert MTB/RIF, TB-RNA and TB-DNA alone and in combination was analyzed. Results The positive detection rates of Xpert MTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid of the smear-negative pulmonary tuberculosis group and the non-tuberculosis pulmonary disease group were 69.96% (354/506) and 2.67% (8/300), 61.46% (311/506) and 5.00% (15/300), and 63.64% (322/506) and 8.00% (24/300), respectively. The rates in the smear-negative pulmonary tuberculosis group were higher than those in the non-tuberculosis lung disease group, and the differences were statistically significant (χ2=342.005, 246.930, 235.687, P<0.01). Compared with the gold standard, the sensitivity, specificity, accuracy, positive predictive value and negative predictive value of Xpert MTB/RIF in the diagnosis of smear-negative pulmonary tuberculosis were 69.96%, 97.33%, 80.15%, 97.79% and 65.77%, respectively; those values of TB-RNA were 61.46%, 95.00%, 73.95%, 95.40% and 59.38%, respectively; those values of TB-DNA were 63.64%, 92.00%, 74.19%, 93.06% and 60.00%, respectively; those values of combined diagnosis with Xpert MTB/RIF, TB-RNA and TB-DNA were 61.26%, 100.00%, 75.68%, 100.00% and 60.48%, respectively; the specificity and positive predictive value of combined detection were higher than those of single detection (P<0.05). Conclusions The joint detection of Xpert MTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid can improve the diagnostic efficacy of smear-negative pulmonary tuberculosis and is worthy of clinical promotion and application.
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The Omicron family of SARS-CoV-2 variants are currently driving the COVID-19 pandemic. Here we analyzed the clinical laboratory test results of 9911 Omicron BA.2.2 sublineages-infected symptomatic patients without earlier infection histories during a SARS-CoV-2 outbreak in Shanghai in spring 2022. Compared to an earlier patient cohort infected by SARS-CoV-2 prototype strains in 2020, BA.2.2 infection led to distinct fluctuations of pathophysiological markers in the peripheral blood. In particular, severe/critical cases of COVID-19 post BA.2.2 infection were associated with less pro-inflammatory macrophage activation and stronger interferon alpha response in the bronchoalveolar microenvironment. Importantly, the abnormal biomarkers were significantly subdued in individuals who had been immunized by 2 or 3 doses of SARS-CoV-2 prototype-inactivated vaccines, supporting the estimation of an overall 96.02% of protection rate against severe/critical disease in the 4854 cases in our BA.2.2 patient cohort with traceable vaccination records. Furthermore, even though age was a critical risk factor of the severity of COVID-19 post BA.2.2 infection, vaccination-elicited protection against severe/critical COVID-19 reached 90.15% in patients aged ≽ 60 years old. Together, our study delineates the pathophysiological features of Omicron BA.2.2 sublineages and demonstrates significant protection conferred by prior prototype-based inactivated vaccines.
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Humans , Aged , Middle Aged , COVID-19/prevention & control , SARS-CoV-2 , Pandemics/prevention & control , China/epidemiology , Disease Outbreaks/prevention & control , VaccinationABSTRACT
@#Abstract: Objective To analyze the clinical characteristics of children with lobar pneumonia and the distribution of pathogens in bronchoalveolar lavage fluid (BALF) collected from these patients, hence providing a scientific basis for their precise diagnosis and treatment. Methods A total of 115 children diagnosed with lobar pneumonia from August 2019 to August 2022 at Suining Central Hospital were screened as the research subjects. The clinical manifestations and occurrence of complications in the patients were investigated. All the children underwent bronchoalveolar lavage after admission, and BALF samples were collected. Fluorescence quantitative PCR was adopted to detect and analyze the distribution and clinical characteristics of Streptococcus pneumoniae (SP) and other related pathogenic microorganisms in BALF specimens. Results Among the 115 pediatric patients with lobar pneumonia, the occurrence of manifestations or complications including involvement of ≥2 lung lobes, myocardial damage, pleural effusion, abnormal liver function, digestive system involvement, nervous system involvement, rash, renal function impairment, and lung atelectasis were observed in 46, 46, 39, 33, 18, 17, 11, 5, and 4 cases, respectively. The pathogen positivity rate in the BALF samples of the 115 patients was 87.0% (100/115), with 81 cases of single infection and 19 cases of mixed infection. A total of 121 strains of pathogens were isolated, including 83 strains of Mycoplasmal pneumonia (MP) (accounting for 68.6%) and SP(13.2%). The differences in the detection rates of HI, MP, RSV strains among different age groups were statistically significant (χ2=8.834, 19.454, 10.284, P<0.05), while the differences in the infection rates of SP, KP, CP, and ADV were not statistically significant (χ2=3.393, 2.67, 0.565, 0.097, P>0.05). The MP pneumonia group showed significantly higher incidence of complications such as pleural effusion, nervous system involvement, and abnormal liver function than the non-MP pneumonia group (χ2=3.925, 4.195, and 4.513, P<0.05). The highest pathogen detection rate was in winter, accounting for 33.91%. Conclusions MP is the most common pathogen in BALF of children with lobar pneumonia. There is variation in the pathogen detection rate among different age groups and seasons. Those with combined infections were more prone to complications, which is worthy of attention by clinicians.
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Objective: To explore the composition of bacteria in lower respiratory tract of patients with pneumoconiosis and dust exposure, and to compare and analyze the difference and correlation between them. Methods: From May 2020 to January 2021, a prospective multicenter cross-sectional study was conducted to select patients with pneumoconiosis who underwent bronchoalveolar lavage treatment at the Respiratory and Critical Care Medical Department of the 920th Hospital of the Joint Support Force and the Respiratory Department of Tongren Hospital in Kunming, as well as the population of dust recipients. A total of 24 patients with pneumoconiosis (pneumoconiosis group) were included, and 16 dust exposed individuals (dust exposed group) were used as controls. Two groups of patients' alveolar lavage fluid were collected. The 16SrRNA gene V3-V4 sequencing technology and bioinformatics analysis platform were used to measure and analyze the differences in microbial structure composition and associations between bacterial communities. Results: Compared with the dust exposed group, the top 5 bacterial phyla in the alveolar lavage fluid level of patients with pneumoconiosis were the same, followed by Proteobacteria, Firmicutes, Bacteroidetes, Fusobacteria, and Actinobacteria. Compared with the dust exposure group, the pneumoconiosis group patients belong to the top 5 genera of horizontal flora abundance, which are different. The dust exposure group is respectively: Pseudomonas, Proctor, Streptococcus, Achromobacter, and Neisseria. The pneumoconiosis group is respectively: Pseudomonas, Achromobacter, Streptococcus, Ralstonia, and Proctor. The Alpha diversity analysis results showed that compared with the dust exposed group, the level of bacterial diversity in the pneumoconiosis group was difference (P<0.05), and there was no statistically significant difference in bacterial evenness (P>0.05) ; Beta diversity showed differences in microbial community structure between the two groups (P<0.05 ). Single factor microbial association network analysis showed that there was a high correlation between Firmicutes and Bacteroidetes in the pneumoconiosis and dust exposed groups and other species, showing a positive correlation; The correlation between Proteobacteria and other species is high, showing a negative correlation. Conclusion: The structure and relative abundance of bacteria in lower respiratory tract were different between patients with pneumoconiosis and dust exposure, and the diversity of bacteria in lower respiratory tract increased in patients with pneumoconiosis, which may be related to disease status.
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Humans , Cross-Sectional Studies , Prospective Studies , Pneumoconiosis , Bacteria/genetics , Dust , Respiratory SystemABSTRACT
OBJECTIVES@#To investigate the clinical application of metagenomic next-generation sequencing (mNGS) of bronchoalveolar lavage fluid (BALF) in the etiological diagnosis and treatment of refractory pneumonia (RTP) in children.@*METHODS@#A retrospective analysis was performed on 160 children with RTP who were admitted to the Department of Pediatric Internal Medicine, Maternal and Child Health Hospital of Inner Mongolia Autonomous Region, from January 2020 to March 2023. According to whether mNGS was performed, they were divided into two groups: mNGS (n=80) and traditional testing (n=80). All children received the tests of inflammatory markers and pathogen tests after admission. Traditional pathogenicity tests included microbial culture (sputum specimen collected by suction tube), nucleic acid detection of respiratory pathogens, and serological test (mycoplasma, tuberculosis, and fungi). For the mNGS group, BALF specimens were collected after bronchoscopy and were sent to the laboratory for mNGS and microbial culture. The two groups were analyzed and compared in terms of the detection of pathogens and treatment.@*RESULTS@#Compared with the traditional testing group, the mNGS group had a significantly higher detection rate of pathogens (92% vs 58%, P<0.05), with more types of pathogens and a higher diagnostic rate of mixed infections. Compared with the traditional testing group, the mNGS group had a significantly higher treatment response rate and a significantly lower incidence rate of complications during hospitalization (P<0.05). Treatment was adjusted for 68 children in the mNGS group according to the results of mNGS, with a treatment response rate of 96% (65/68) after adjustment.@*CONCLUSIONS@#Compared with traditional pathogen tests, BALF mNGS can significantly improve the detection rate of pathogens and find some rare pathogens. In clinical practice, when encountering bottlenecks during the diagnosis and treatment of children with RTP, it is advisable to promptly perform the mNGS to identify the pathogens.
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Humans , Child , Bronchoalveolar Lavage Fluid , Retrospective Studies , Pneumonia/therapy , High-Throughput Nucleotide Sequencing , Bronchoscopy , Sensitivity and SpecificityABSTRACT
Objective:To investigate drug resistance gene in Mycoplasma pneumoniae(MP) and the distribution of 13 respiratory pathogens in bronchoalveolar lavage fluid(BALF) of children with Mycoplasma pneumoniae pneumonia(MPP).Methods:A total of 100 BALF of children with MPP in Peking University Third Hospital and Peking University First Hospital from January 2018 to January 2019 were collected.Fluorogenic quantitative PCR was used to detect nucleic acid and it′s drug resistance gene of MP and multiple PCR method was adopted to detect influenza A virus, influenza A virus-H 1N 1, influenza A virus-H 3N 2, influenza B, human parainfluenza virus, adenovirus, human bocavirus, human rhinovirus, Chlamydia pneumoniae, human metapneumovirus, MP, human coronavirus, and respi-ratory syncytial virus gene, and the results were compared by using Chi square test. Results:In 100 BALF samples, MP and drug resistance gene were detected by fluorogenic quantitative PCR.Totally, 83 cases (83.00%) were MP positive and 78 cases (93.98%) were drug resistant.All of them had the point mutations A2063G in V region of 23S rRNA domain.A total of 13 kinds of respiratory pathogens were detected by multiplex PCR method, and 89 cases (89.00%) were positive.Totally, 79 cases (79.00%) were MP positive, of which 74 cases (74.00%) detected only MP, and 5 cases (5.00%) detected MP combined with other pathogens.Other pathogens were detected in 10 cases (10.00%). The virus detection rate of 0-4 years old group was higher than that of >4-6 years old group ( P=0.042) and >6 years old group ( P=0.002), and the differences were statistically significant. Conclusions:MP can be detected in most BALF samples of MPP children, the drug resistance phenomenon is serious, and the main point mutation is A2063G.There were other respiratory pathogens and 2 or 3 pathogens were detected in a small number of BALF samples.
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Objective:To investigate clinical and bronchoscopy characteristics of Mycoplasma pneumoniae pneumonia in children with 23S rRNA resistance gene positive in bronchoalveolar lavage fluid(BALF), and find clinical indicators that can identify Mycoplasma pneumoniae drug resistance early.Methods:The clinical data of 61 hospita-lized children diagnosed with Mycoplasma pneumoniae pneumonia as subjects from October 2017 to June 2018 in the Department of Pediatric Respiratory Medicine of Shengjing Hospital Affiliated to China Medical University were analyzed retrospectively.Bronchoscopy was performed on each subject and the BALF was taken and used to detect the 2063 site mutation of the 23S rRNA V region gene in BALF, and they were divided into drug-resistant gene positive group and drug-resistant gene negative group.The clinical manifestations, relevant laboratory data, imaging data, and bronchoscopy findings in different load groups were compared.Statistical methods such as t test, rank sum test, χ2 test, Fisher′ s exact probability method, and multivariate Logistic regression analysis were used to analyze the data. Results:Among the 61 children, 38 cases (62.30%) were in the drug resistance gene positive group, and 23 cases (37.70%) were in the drug resistance gene negative group.There were no significant differences in gender and age between the two groups (all P>0.05). The days of hospitalization and fever in the children with positive drug resistance genes were longer than those in the negative drug resistance gene groups, and they were more likely to have refractory mycoplasma pneumoniae pneumonia(RMPP) and extra pulmonary complications, with statistically significant differences ( P<0.05), but there were no significant differences in hypoxemia ( P>0.05). There were significant differences in white blood cell(WBC), C-reactive protein(CRP), procalcitonin(PCT), D-Dimer (D-D) and interleukin-6 (IL-6) between the two groups (all P<0.05). Except that the WBC level in the drug-resistant gene-positive group was lower than that in the drug-resistant gene-negative group, the rest of the test results indicated that the drug-resistant gene-positive group was higher than the drug-resistant gene-negative group.There were no significant differences in the concentrations of serum lactate dehydrogenlase(LDH)and IL-6 in BALF ( P>0.05). In this study, Logistic regression analysis was performed on several statistically significant laboratory indicators.It was found that WBC was more sensitive to identify drug resistance genes, and the optimal critical value was 8.55×10 9/L.The specificity of D-D in identifying drug resistance genes was higher, and the optimal cut-off value was 523 μg/L.In the drug resistance gene positive group, 35 cases (92.11%) showed extensive lung consolidation/atelectasis on imaging, and the drug resistance gene negative group was 13 cases (56.52%), with statistically significant differences ( P<0.05). The drug resistance gene-positive group mainly showed mucosal erosion, necrosis, phlegm plug/plastic phlegm plug and bronchitis stenosis, with a total of 19 cases (50.00%). In the drug resistance gene negative group, the main manifestations of mucosal longitudinal wrinkle, flocculent and viscous secretions were 14 cases (60.88%), with statistically significant differences ( P<0.05). Conclusions:The point mutation of 23S rRNA V region gene is closely related to the clinical characteristics of children with Mycoplasma pneumoniae pneumonia.Children with A2063G mutation are more prone to have RMPP and extrapulmonary complications, and their imaging manifestation and bronchoscopy are more severe.The levels of leukocytes and D-D in the blood have significance for the early identification of drug resistance.The systemic excessive immune inflammatory response caused by Mycoplasma pneumoniae pneumonia with drug-resistant gene positive needs to be valued.
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Objective:To investigate the value of the level of heparin-binding protein(HBP)in bronchoalveolar lavage fluid(BALF)on the evaluation of severe pneumonia in children.Methods:A total of 94 children with severe pneumonia who underwent bronchoscopy and bronchoalveolar lavage were admitted at Hunan Children′s Hospital, and HBP levels in BALF were detected.According to the etiological results, the patients were divided into non-bacterial infection group(19 cases) and bacterial infection group(75 cases). According to the existence and severity of acute respiratory distress syndrome (ARDS), the cases were divided into non-ARDS group(65 cases), mild ARDS group(23 cases) and moderate to severe ARDS group(6 cases).Results:The HBP level of BALF in the bacterial infection group was higher than that in the non-bacterial infection group, and the difference was statistically significant[ 20.77(5.90, 73.50)ng/mL vs.5.9(5.90, 7.64)ng/mL, Z=12.500, P<0.001]. The HBP level of BALF in the moderate to severe ARDS group[300.00(169.29, 300.00)ng/mL] was significantly higher than those in the non-ARDS group[11.90(5.90, 36.95)ng/mL] and the mild ARDS group[15.13(7.41, 46.44)ng/mL], and the difference was statistically significant( H=14.718, P=0.001). In predicting the presence of bacterial infection in severe pneumonia, the area under the receiver operating characteristic curves of BALF HBP, serum procalcitonin (PCT) and serum C-reactive protein(CRP) were 0.758, 0.737, and 0.732, respectively.When the optimal truncation values of BALF HBP, serum PCT and serum CRP were 8.40 ng/mL, 0.16 ng/mL, and 8.39 mg/L, the predicted sensitivities were 70.7%, 69.3%, 46.7%, and the predicted specificity were 79.0%, 79.0%, 94.7%, respectively. Conclusion:The level of HBP in BALF in children with severe pneumonia increases with the severity of ARDS, and significantly increases in the positive group of bacterial infection, which can be used as one of the auxiliary indicators to evaluate the severity of severe pneumonia and bacterial infection in children.
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Objective:To investigate the clinical features, therapy and prognosis of human cytomegalovirus(HCMV)pneumonia in pediatric patients, and to analyze the diagnosis value of detecting HCMV DNA in bronchoalveolar lavage fluid(BALF)by real-time PCR.Methods:The clinical characteristics of 58 pediatric inpatients who were HCMV DNA positive in BALF were retrospectively reviewed.All the patients were from Shengjing Hospital of China Medical University from January 2015 to December 2019.Clinical, radiologic, laboratory and microbiologic data was collected for each patient.The study cohort was divided into HCMV productive infection and latent infection consisting of 22 and 36 patients respectively, based on the HCMV active infection in lung or not.Receiver operating characteristic(ROC)curve was used to assess utility of detecting HCMV DNA in BALF and establish a threshold for diagnosis.Results:(1)Compared with patients in latent infection group, the children in productive infection group had a lower age of onset( P<0.05), a higher proportion of male( P<0.05), and more prolonged hospitalization stay( P<0.05). Pulmonary rales, hypoxemia and higher AST, CK, LDH in serum were easier to detect in productive infection group( P<0.05). Higher HCMV DNA copies in BALF was also detected( P<0.01). Patients in productive infection group had significantly more exposure to additional oxygen treatment or mechanical ventilation and systemic hormone therapy( P<0.05), while with poorer outcomes( P<0.05). (2) ROC curve analysis showed that the AUC for HCMV DNA in BALF in diagnosis of HCMV pneumonia was 0.708 with a threshold of 8.83×10 3 copies/mL, a sensitivity of 77.27%, and a specificity of 58.33%. Conclusion:Those who are diagnosed HCMV pneumonia have a lower age of onset with higher male proportion.These children suffered severer clinical signs.The patients with HCMV DNA copies higher than 8.83×10 3 copies/mL in BALF would be more likely to be diagnosed as HCMV pneumonia.
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Objective:To analyze the distribution and drug sensitivity of pathogens in bronchoalveolar lavage fluid(BALF)of children with severe community acquired pneumonia(CAP)in Qingdao from 2018 to 2020.Methods:The clinical data of 482 children with severe CAP in Qingdao admitted to Women and Children′s Hospital of Qingdao University were collected.BALF was collected by bronchoscopy for detection of bacteria and mycoplasma.Results:(1)Bacterial infection was detected in 139 cases(27.84%), mycoplasma infection in 119 cases(24.69%), and virus infection in 141 cases(29.25%). (2)The detection rates of bacteria and virus infection in the 1-12 months old group were higher.The detection rate of mycoplasma pneumoniae was the highest in the group over 5 years old.(3)A total of 139 strains were positive in bacterial culture of lavage fluid under bronchoscope: 55 strains(39.57%) of gram-negative bacilli and 84 strains(60.43%) of gram-positive cocci.Streptococcus pneumoniae was the most common gram-positive bacteria.Haemophilus influenzae was the most common gram-negative strain.(4)Streptococcus pneumoniae and Staphylococcus aureus were highly sensitive to amoxicillin clavulanate potassium, vancomycin and linezolid.The resistance rate to erythromycin was high(100%). (5)Haemophilus influenzae, Escherichia coli, Pseudomonas aeruginosa and Klebsiella pneumoniae were highly sensitive to meropenem and cefoperazone sulbactam.They were highly resistant to amoxicillin, ampicillin and cefuroxime(>80%).Conclusion:Severe CAP in Qingdao area is mainly caused by virus and bacteria within 1 year old.Mycoplasma pneumoniae infection is the main cause of children over 5 years old.Respiratory syncytial virus, adenovirus and parainfluenza virus are main causes of virus infection.Streptococcus pneumoniae and haemophilus influenzae are the main pathogens, which are more sensitive to vancomycin, linezolid, meropenem and cefoperazone sulbactam, but resistant to erythromycin and amoxicillin.
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Objective@#To assess the effects of acute exposure to electronic cigarette ( e-cigarette ) on leukocyte and total protein levels in bronchoalveolar lavage fluid ( BALF ) and pulmonary surfactant protein expression in a mouse model, so as to provide insights into the elucidation of the mechanism underlying the damages to the respiratory system caused by e-cigarette.@*Methods@#Twenty-one C57BL/6N female mice were randomly divided into the blank control group, the solvent control group and the nicotine group. Mice in the solvent control group and the nicotine group were exposed to the solvent aerosol or e-cigarette aerosol containing 25 mg/mL nicotine for 3 hours daily, while mice in the blank control group were bred in clean air. Following 3-day exposure, mouse BALF and lung specimens were collected. The cell morphology was observed using microscopy following Wright-Giemsa staining and the leukocyte count was estimated in BALF, while the total protein expression was quantified using bicinchoninic acid ( BCA ) assay. In addition, the mRNA expression of pulmonary surfactant protein genes was detected in mouse lung specimens using quantitative real-time PCR ( qPCR ) assay.@*Results@#All mice in three groups grew well without obvious abnormality or death seen. Wright-Giemsa staining showed a higher number of mononuclear macrophages in mouse BALF in the nicotine group than in the blank control group and the solvent control group. The leukocyte counts were ( 2.00±0.77 )×107, ( 1.79±0.99 )×107 and ( 4.00±1.35 )×107 cells/L ( F=9.199, P=0.002 ), and the total protein levels were ( 0.16±0.03 ), ( 0.12±0.02 ) and ( 0.16±0.04 ) mg/mL in mouse BALF in the blank control group, solvent control group and nicotine group ( F=3.610, P=0.048 ), and the relative mRNA expression of pulmonary surfactant protein B (SP-B) and SP-D was 1.00±0.14, 0.82±0.12 and 0.74±0.07 ( F=5.491, P=0.028 ), and 1.00±0.06, 0.90±0.02 and 0.71±0.15 in mouse lung specimens, respectively ( F=10.460, P=0.005 ). The leukocyte count was significantly higher in the nicotine group than in the blank control group and solvent control group (P=0.007, 0.003), and the total protein content was higher in the nicotine group than in the solvent control group ( P=0.060 ), while the relative SP-B mRNA expression was lower in the nicotine group than in the blank control group ( P=0.025 ), and the relative SP-D mRNA expression was lower in the nicotine group than in the blank control group and solvent control group ( P=0.004, 0.041 ).@*Conclusion@#Acute exposure to e-cigarette results in elevated intrapulmonary inflammatory responses, pulmonary capillary barrier impairment and reduced pulmonary surfactant protein expression.
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Objective:To investigate the effect of empirical antifungal treatment on the diagnostic sensitivity of galactomannan(GM)in bronchoalveolar lavage(BALF)in chronic obstructive pulmonary disease(COPD)patients combined with invasive pulmonary aspergillosis(IPA).Methods:COPD patients with IPA were enrolled between January 2015 and January 2019 as research objects.Patients who were treated with antifungal drugs prior to bronchoscopy were considered as the empirical group, and other patients were considered as the diagnosis-driven group.The results of BALF GM were compared between the two groups.Results:A total of 66 COPD patients with IPA were enrolled in this study, with 5 cases confirmed and 61 cases clinically diagnosed.Of them, 17 cases were in the empirical group and 49 in the diagnosis-driven group.There was no significant difference in the sensitivity of blood GM and microbiological examination between the two groups( χ2=0.248 and 0.379, P=0.619 and 0.538). With BALF GM 0.5 as a cutoff value, the sensitivity of BALF GM was slightly lower in the empirical group than in the diagnosis-driven group but with no significant difference(88.2%, 95% CI: 62.3%-97.9% vs.93.9%, 95% CI: 82.1%-98.4%, χ2=0.051, P=0.821). However, with BALF GM 1.0 as a cutoff value, the sensitivity decreased greatly in the empirical group compared with the diagnosis-driven group(52.9%, 95% CI: 28.5%-76.1% vs.80.6%, 95% CI: 67.4%-90.8%, χ2=4.036, P=0.045). Logistic regression analysis showed that after adjusting for mechanical ventilation( OR=0.807, 95% CI: 0.215-3.026, P=0.750)and use of semisynthetic penicillins( OR=0.498, 95% CI: 0.140-1.776, P=0.283), the false negative rate of BALF GM was associated with empirical antifungal therapy( OR=0.243, 95% CI: 0.068-0.949, P=0.030). Conclusions:Empirical antifungal therapy prior to bronchoscopy can decrease the diagnostic sensitivity of BALF GM in COPD patients with IPA.
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Objective:To analyze the risk factors,clinical characteristics and prognosis of the pneumocystis pneumonia(PCP) that is one of the severe pulmonary complications after allogeneic hematopoietic stem cell transplantation(allo-HSCT).Methods:The clinical features,laboratory data,treatment and outcomes of patients with PCP after allo-HSCT in our hospital from January,2016 to January,2021 were retrospectively collected and analyzed.Results:Twenty three cases who met the clinical diagnostic criteria of PCP were enrolled. The median time of diagnosed as PCP after transplantation was 221 days. The computed tomography (CT) of chest indicated diffuse ground glass opacity.The median of β-1,3-D glucan consentration was 894.25 ng/L, and 91.3% of the cases were over 60 ng/L.The lymphocyte count in 60.9% cases was lower than 1×10 9/L;CD4 +T lymphocyte count in 65.2% of patients was less than 200/μL. Pneumocytis sequences of mNGS were positive in all 21 cases.15 patients were complicated with mixed infection.All patients were treated with TMP-SMX,18 patients were cured and 5 patients died. Conclusions:Patients with PCP after allo-HSCT progresses rapidly, and which is usually with multiple infections. Serum β-1,3-D glucan concentration increase contributes to the diagnosis of PCP.And mNGS in alveolar lavage fluid is highly sensitive to Pneumocystis, which helps patients get treatment in time, so as to reduce mortality.Patients with respiratory failure progressing to a need for mechanical ventilation and high flow oxygen inhalation suggest a poor prognosis.
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Objective To evaluate the diagnostic value of quantitative detection of cytomegalovirus (CMV) DNA from different sources [plasma, sputum and bronchoalveolar lavage fluid(BALF)] for CMV pneumonia after allogeneic hematopoietic stem cell transplantation. Methods Clinical data of 405 recipients undergoing allogeneic hematopoietic stem cell transplantation were retrospectively analyzed. Among them, 19 recipients diagnosed with CMV pneumonia were assigned into the CMV pneumonia group, and 229 recipients with CMV viremia alone, 11 recipients without CMV pneumonia who received fiberoptic bronchoscopy and 16 recipients diagnosed with bacterial or fungal pneumonia based on pathogenic evidence receiving sputum culture were assigned into the control A, B and C groups, respectively. The incidence of CMV pneumonia was summarized. The CMV DNA load of specimens from different sources (plasma, sputum and BALF) of recipients with CMV pneumonia was analyzed. The clinical prognosis of recipients with CMV pneumonia was evaluated. Results Among 405 recipients undergoing allogeneic hematopoietic stem cell transplantation, 19 cases developed CMV pneumonia, and the overall incidence of CMV pneumonia was 4.7%(19/405). The CMV DNA load in the plasma, sputum and BALF of recipients with CMV pneumonia was higher than those in the control A, B and C groups (all P < 0.05). In the 19 recipients, 12 cases were cured after antiviral treatment and 7 died from treatment failure(3 cases abandoned treatment). The fatality was 37%(7/19). Conclusions Quantitative detection of CMV DNA in the plasma, sputum and BALF may increase the diagnostic rate of CMV pneumonia, thereby improving clinical prognosis of recipients undergoing allogeneic hematopoietic stem cell transplantation.
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Objective:To evaluate the diagnostic value of metagenomics next-generation sequencing (mNGS) in detecting pathogens in bronchoalveolar lavage fluid (BALF) for pulmonary infection in solid organ transplant patients in intensive care unit (ICU).Methods:A retrospective study was conducted, the BALF samples from 46 patients with post organ transplant pneumonia/suspected pneumonia admitted to the Department of Critical Care Medicine of the First Affiliated Hospital of University of Science and Technology of China from August 2018 to August 2021 were collected, all tested by simultaneous mNGS and conventional comprehensive microbial test (CMT), and the results of CMT were used as the reference standard to compare the differences in the diagnostic value of mNGS and CMT for pulmonary infections in solid organ transplant patients, and to analyze the diagnostic value of mNGS for mixed infections.Results:① Pneumonia pathogens: a total of 31 pathogens were detected in 35 patients, including bacteria (16 species), fungi (9 species) and viruses (6 species). Among them, 25 pathogens were detected by mNGS and CMT, and only 19 pathogens were detected by mNGS. Among the microorganisms isolated by mNGS method, the detection rates of Pseudomonas aeruginosa, Acinetobacter baumannii and Klebsiella pneumoniae were higher [51.4%(18/35), 42.9% (15/35), 31.4% (11/35), respectively]; Candida albicans, Aspergillus and Pneumocystis carinii were the most commonly detected fungi [31.4% (11/35), 22.9% (8/35), 22.9% (8/35), respectively]; 20 patients were positive for the virus, and the most commonly detected viruses were cytomegalovirus, herpesvirus and EB virus [28.6% (10/35), 20.0% (7/35), 17.1% (6/35), respectively]. In addition, one case of Brucella was detected by mNGS.② Diagnostic efficiency: as far as bacterial detection is concerned, 20 cases of negative results were obtained by CMT detection of 35 samples included in the study, and a total of 10 cases of positive results were obtained by mNGS detection of negative samples; the percentage of mNGS positive samples was significantly higher than that of CMT positive samples [odds ratio ( OR) = 5.5, 95% confidence interval (95% CI) = 1.2-24.8, P = 0.02]. When compared with CMT, the sensitivity and specificity of mNGS were 93.3% and 50.0%, and the positive predictive value (PPV) and negative predictive value (NPV) were 58.3%, 91.1%. As far as fungal detection was concerned, there was no significant difference in the percentage of positive samples between the two methods ( OR = 1.5, 95% CI = 0.5-4.2, P = 0.60); the sensitivity and specificity of mNGS were 72.2% and 64.7%, and the PPV and NPV were 68.4%, 68.8%; CMT test of the 35 included samples produced 17 negative results, and mNGS test of the negative samples produced 6 positive results. A total of 20 patients tested positive for the virus by mNGS. In addition, 23 patients (65.7%) were diagnosed with pulmonary mixed infection. Conclusion:The use of mNGS to detect pathogens in BALF can improve the sensitivity and specificity of bacterial identification of pulmonary infection in critically ill organ transplant patients, and mNGS has obvious advantages in detecting virus and identifying mixed infections.