Evaluation of respiratory samples in etiology diagnosis and microbiome characterization by metagenomic sequencing.

BACKGROUND: The application of clinical mNGS for diagnosing respiratory infections improves etiology diagnosis, however at the same time, it brings new challenges as an unbiased sequencing method informing all identified microbiomes in the specimen. METHODS: Strategy evaluation and metagenomic analysis were performed for the mNGS data generated between March 2017 and October 2019. Diagnostic strengths of four specimen types were assessed to pinpoint the more appropriate type for mNGS diagnosis of respiratory infections. Microbiome complexity was revealed between patient cohorts and infection types. A bioinformatic pipeline resembling diagnosis results was built based upon multiple bioinformatic parameters. RESULTS: The positive predictive values (PPVs) for mNGS diagnosing of non-mycobacterium, Nontuberculous Mycobacteria (NTM), and Aspergillus were obviously higher in bronchoalveolar lavage fluid (BALF) demonstrating the potency of BALF in mNGS diagnosis. Lung tissues and sputum were acceptable for diagnosis of the Mycobacterium tuberculosis (MTB) infections. Interestingly, significant taxonomy differences were identified in sufficient BALF specimens, and unique bacteriome and virome compositions were found in the BALF specimens of tumor patients. Our pipeline showed comparative diagnostic strength with the clinical microbiological diagnosis. CONCLUSIONS: To achieve reliable mNGS diagnosis result, BALF specimens for suspicious common infections, and lung tissues and sputum for doubtful MTB infections are recommended to avoid the false results given by the complexed respiratory microbiomes. Our developed bioinformatic pipeline successful helps mNGS data interpretation and reduces manual corrections for etiology diagnosis.

The clinical value of valve metagenomic next-generation sequencing when applied to the etiological diagnosis of infective endocarditis.

BACKGROUND: Metagenomic next-generation sequencing (mNGS) is widely applied in the etiological diagnosis of infectious diseases. However, the clinical practice of mNGS in infective endocarditis (IE) is relatively less studied. This research aimed to assess the etiological diagnostic value of valve mNGS in IE. METHODS: We retrospectively analyzed 49 IE patients who underwent cardiac valve surgery in Zhongshan Hospital, Fudan University, Shanghai from 1 June 2018 to 30 November 2020. Among these IE patients, 28 were culture positive and 21 were culture negative. The culture results of the culture-positive IE patients were set as gold standard to assess the sensitivity and specificity of valve mNGS in the etiological diagnosis of IE. We studied the positive detection rate of pathogens by valve mNGS among the culture-negative IE patients. During the same period, we also collected the resected valves of 8 patients with non-infective valvular diseases for mNGS as negative controls. RESULTS: The valve mNGS results of the culture-positive IE patients were the exact same as their culture results. Both the sensitivity and specificity of valve mNGS were 100%. The positive detection rate of pathogens by valve mNGS was 100% among the culture-negative IE patients. The stringent mapped reads number of genera (SMRNG), relative abundance of genera, stringent mapped reads number of species (SMRN), relative abundance of species, and coverage rate of valve mNGS results were significantly higher in culture-positive IE participants than in culture-negative IE participants. The valve mNGS results of the 8 participants with non-infective valvular diseases were all negative. CONCLUSIONS: Valve mNGS is a promising technology for the etiological diagnosis of IE, especially culture-negative IE, and it may be used to guide precise antibiotic treatment after surgery.

Identification of metabolites of the novel anti-tumor drug candidate MDH-7 in rat urine by liquid chromatography coupled with triple quadrupole linear ion trap mass spectrometry.

RATIONALE: Our previous preliminary pharmacokinetic study demonstrated that the novel double pyrimidine tricyclic nucleoside MDH-7 in rats had a very short half-life (<30 min) after oral administration. As a result, the in vivo metabolic profile of MDH-7 should be investigated during early stages of drug development to better select drug candidates. METHODS: In this study, a rapid method was developed to identify the metabolites of MDH-7 in rat urine by means of ultra-performance liquid chromatography (UPLC) coupled with electrospray ionization mass spectrometry (ESI-MS) using a triple quadrupole linear ion trap instrument. MDH-7 and its metabolites were detected and characterized by the combined use of the multiple reaction monitoring-information-dependent acquisition-enhanced product ion (MRM-IDA-EPI) mode and the precursor scan information-dependent acquisition-enhanced product ion (PREC-IDA-EPI) mode. RESULTS: Ten novel metabolites of MDH-7 were identified and characterized in rat urine by LC/ESI-MS and collision-induced dissociation tandem mass spectrometry (CID-MS/MS) analyses. M1 was identified as 5-fluoro-N(4) -[(pentyloxy)carbonyl]cytosine; M2 and M3 were formed by hydroxylation products of M1. Metabolites M4-M10 were formed by a series of degradation reactions such as: deacetylation, hydroxylation, loss of the defluorocytosine base, oxidative-deamination, loss of the defluorouracil base, N-dealkylation and amide hydrolysis. CONCLUSIONS: Based on the profiles of the metabolites, possible metabolic pathways of MDH-7 in rats were proposed for the first time. This study provides new and available information on the metabolism of MDH-7 which is very useful to further understand its in vivo metabolic fate. Copyright © 2016 John Wiley & Sons, Ltd.

[The evaluation of strengthened psychological and behavioral intervention in smoking cessation clinics].

OBJECTIVE: To evaluate the effect of psychological and behavioral intervention combined with varenicline smoking cessation clinics and to analyze predictors of successful quitting. METHODS: Subjects were collected from quitters who went to receive consultation and intervention in "smoking and related diseases" clinic at Zhongshan Hospital, Fudan University from March 2009 to September 2010. Eligible subjects were screened and divided into strengthen follow-up group and control group. The 4 weeks continuous abstinence rate from week 9 through week 12 were observed logistic regression model and used to analyze the predictors of successful quitting. RESULTS: Subjects who are addicted to nicotine received strengthening psychological and behavioral intervention combined with varenicline in smoking cessation clinics. The total continuous cessation rate during the 9(th) - 12(th) week was 52.3%, with 60.9% (28/46) and 46.2% (30/65) of strengthen follow-up group and control group respectively. The most frequent adverse effects were nausea 39.6% (44/111), insomnia and abnormal dreams 17.1% (19/111). Adverse effects were tolerable and withdraw symptoms were few. Preparation and medication time can be used as predictors of successful quitting. CONCLUSION: The quit rate of varenicline therapy combining with strengthen intervention is high and strengthening psychological and behavioral intervention could increase the success rate more obviously, which is a good choice for cessation therapy in smoking cessation clinics. Better preparation and regular adequate treatment can improve quit rate.