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1.
Infect Drug Resist ; 17: 1625-1632, 2024.
Article in English | MEDLINE | ID: mdl-38699076

ABSTRACT

Objective: Exploring the risk factors for readmission of elderly patients with pulmonary tuberculosis (PTB) within one month using the propensity score matching(PSM). Methods: A retrospective analysis was conducted on the clinical data of elderly patients with PTB who were admitted to the Tuberculosis Department of Lishui Hospital of Traditional Chinese Medicine from January 2020 to October 2023. The patients were divided into two groups: non-readmission group and readmission group based on whether they were readmitted within one month after discharge. The PSM method was used to match the baseline data of the two groups of patients, and multivariate logistic regression analysis was conducted to explore the risk factors for readmission of elderly patients with PTB within one month after discharge. Results: A total of 1268 hospitalized elderly patients with PTB were included in the study, comprising 977 readmitted patients and 291 newly admitted patients (22.95%). Using the PSM, 288 pairs of patients were successfully matched. Following matching, there were no statistically significant differences between the two groups in terms of gender, age, occupation, body mass index(BMI), past medical history, etc. (all P>0.05). Multivariate logistic regression analysis indicated that infection, drug-induced liver injury(DILI), acute heart failure(AHF), chronic kidney disease(CKD), and extrapulmonary tuberculosis(EPTB) were all identified as risk factors for readmission of elderly patients with PTB. Conclusion: After controlling for confounding factors through PSM, the study revealed that infection, DILI, AHF, CKD, and EPTB are risk factors for readmission among elderly patients with PTB, highlighting the need for early intervention.

2.
J Ethnopharmacol ; 329: 118142, 2024 Jul 15.
Article in English | MEDLINE | ID: mdl-38583730

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: Psoralea corylifolia L. (Fabaceae) is a traditional medicinal herb used to treat various diseases, including kidney disease, asthma, psoriasis and vitiligo. AIM OF THE STUDY: To explore the antibacterial activity of Psoralea corylifolia L. and its bioactive components against Mycobacterium abscessus (M. abscessus). MATERIALS AND METHODS: Ultra high performance liquid chromatography was utilized to analyze the bioactive fractions and compounds present in 30%, 60%, and 90% ethanol extracts of Psoralea corylifolia L.. The antibacterial effects of Psoralea corylifolia L. and potential active ingredients were determined by minimum inhibitory concentration (MIC). The bactericidal activity of the active ingredient isobavachalcone was evaluated and then scanning electron microscopy was used to explore the bactericidal mechanism of isobavachalcone. RESULTS: The 90% ethanol extracts of Psoralea corylifolia L. showed significant antibacterial activity against M. abscessus, with an MIC of 156 µg/mL. Isobavachalcone was identified as the bioactive ingredient, and testing of 118 clinical isolates of M. abscessus indicated their MICs ranged from 2 to 16 µg/mL, with an average MIC of 8 µg/mL. Furthermore, the minimum bactericidal concentration/MIC ratio and the time-kill test indicated rapid bactericidal activity of isobavachalcone against M. abscessus. Finally, we found that the bactericidal mechanism of isobavachalcone involved damage to the bacterial cell membrane, causing wrinkled and sunken cell surface and a noticeable reduction in bacterial length. CONCLUSION: Psoralea corylifolia L. ethanol extracts as well as its active component isobavachalcone show promising antimicrobial activity against M. abscessus.


Subject(s)
Anti-Bacterial Agents , Chalcones , Microbial Sensitivity Tests , Mycobacterium abscessus , Plant Extracts , Psoralea , Psoralea/chemistry , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/isolation & purification , Anti-Bacterial Agents/chemistry , Plant Extracts/pharmacology , Plant Extracts/chemistry , Chalcones/pharmacology , Chalcones/isolation & purification , Mycobacterium abscessus/drug effects
3.
Infect Drug Resist ; 17: 275-282, 2024.
Article in English | MEDLINE | ID: mdl-38298533

ABSTRACT

Purpose: Sangju-Yin, supplemented with some drugs, has frequently demonstrated therapeutic efficacy against colds, albeit its effect on Mycoplasma pneumoniae (MP) infection remains unknown. Therefore, we aimed to elucidate the treatment efficacy and influencing factors of a Sangju cough mixture on MP infection in adults. Patients and Methods: Between January 2021 and December 2022, 150 adult patients with MP infection at the Lishui Hospital of Traditional Chinese Medicine Affiliated with Zhejiang University of Traditional Chinese Medicine were assigned to the treatment (administered Sangju cough mixture and moxifloxacin tablets) or the control (administered moxifloxacin tablets) groups. Results: When compared with the control group, the treatment group exhibited significantly improved traditional Chinese medicine syndrome scores, increased CD4+ T cell levels, and decreased CD8+ T cell levels (all P < 0.05). After 7 days of treatment, the negative conversion rate of the MP-specific immunoglobulin M (MP-IgM) antibody of the treatment group was not significantly different from that of the control group (P > 0.05); however, after 14 days of treatment, the rate was significantly higher in the treatment group (P < 0.05). The univariate regression analysis revealed that combined chronic respiratory disease, failure to take Sangju cough mixture, combined pneumonia, Nutritional Risk Screening 2002 (NRS 2002) score of at least 3 points, and age were associated with the negative conversion of the MP-IgM antibody (all P < 0.05). Nevertheless, the multivariate regression model revealed that the NRS 2002 score of at least 3 points was not an independent risk factor (P > 0.05). Conclusion: Sangju cough mixture can improve symptoms, accelerate the negative conversion time of MP-IgM antibody, and promote rehabilitation of the patients.

4.
Patient Prefer Adherence ; 18: 227-237, 2024.
Article in English | MEDLINE | ID: mdl-38274356

ABSTRACT

Objective: The purpose of this study is to examine the effect of quality control circle (QCC) activities on increasing the mask-wearing compliance of hospitalized patients diagnosed with tuberculosis (TB) during external examinations. Methods: To assess the mask-wearing compliance of patients diagnosed with TB admitted to a ward in our hospital in 2019, who visited other departments, we conducted activities in accordance with the ten steps of the QCC. We outlined the causes of non-compliance and developed and implemented improvement plans. We compared the results obtained before and after the implementation of the QCC to assess the enhancement in patient compliance concerning wearing masks during external examinations, particularly among individuals diagnosed with pulmonary tuberculosis. Results: The compliance rate for mask-wearing during external examinations rose from an initial 32.61% (45/138) prior to the intervention to 83.71% (149/178) following the intervention. The difference between the two groups was statistically significant (χ2 = 85.635, P < 0.001). The following countermeasures and implementation measures were formulated: (1) Increasing public health education and awareness; (2) Strengthening the training of nursing staff to enhance their knowledge; (3) Providing free surgical masks; (4) Establishing an effective monitoring system with the department of auxiliary examinations. Conclusion: QCC interventions were significantly effective in enhancing adherence to mask-wearing protocols during external examinations of hospitalized patients with tuberculosis. Such improvements contribute substantially to the mitigation of tuberculosis transmission within clinical and healthcare environments.

5.
BMC Pulm Med ; 23(1): 451, 2023 Nov 20.
Article in English | MEDLINE | ID: mdl-37986162

ABSTRACT

OBJECTIVE: The objective of this study is to analyze the risk factors associated with bronchiectasis combined with non-tuberculous mycobacteria pulmonary disease(NTM-PD) and provide a basis for more effective prevention and treatment strategies. METHODS: The study subjects for this manuscript were patients with bronchiectasis who were admitted to the infection department between January 2021 and June 2023.There were 34 patients with NTM-PD in the observation group, and 52 patients with simple bronchiectasis in the control group. Basic information, imaging features, serum albumin levels, and infection indicators were collected from both groups of patients.Univariate and multivariate logistic regression analysis were performed to analyze the risk factors for NTM-PD in patients with bronchiectasis. RESULTS: Multivariate logistic regression analysis revealed that bronchiectasis exacerbation occurring at least twice a year(OR = 3.884, 95% CI: 1.200-12.568), involvement of three or more lung lobes with bronchiectasis (OR = 3.932, 95% CI: 1.208-12.800), hypoalbuminemia (OR = 3.221, 95% CI: 1.015-10.219), and the NLR index (OR = 1.595, 95% CI: 1.200-2.119) were significant risk factors for non-tuberculous mycobacteria pulmonary disease in individuals with bronchiectasis (P < 0.05). CONCLUSION: Patients with bronchiectasis accompanied by NTM-PD present specific risk factors that should be promptly addressed through prevention and treatment.


Subject(s)
Bronchiectasis , Lung Diseases , Mycobacterium Infections, Nontuberculous , Humans , Nontuberculous Mycobacteria , Mycobacterium Infections, Nontuberculous/complications , Mycobacterium Infections, Nontuberculous/epidemiology , Mycobacterium Infections, Nontuberculous/microbiology , Case-Control Studies , Bronchiectasis/complications , Bronchiectasis/epidemiology , Bronchiectasis/microbiology , Lung Diseases/epidemiology , Lung Diseases/complications , Risk Factors , Retrospective Studies
6.
Chem Biodivers ; 20(12): e202301345, 2023 Dec.
Article in English | MEDLINE | ID: mdl-37985414

ABSTRACT

Marine actinomycetes are known for their production of remarkable organic molecules, particularly those featuring polyoxygenated long-chain backbones. Determining the absolute configurations of these compounds remains a challenging task even today. In this study, we successfully established the planar structures and absolute configurations of two highly flexible amide alkaloids from Streptomyces sp. WU20: kueishanamides A (1) and B (2). These compounds possess a C13 linear backbone and each contains five stereogenic carbon centers. Our approach involved a combination of spectroscopic and computational methods, including J-based configurational analysis and VCD calculations, ensuring the unambiguous determination of their configurations. Kueishanamide A (1) and kueishanamide B (2) showed moderate antifungal activity against pathogenic fungus Crytococcus neoformans, with MIC values of 25 µg/mL each.


Subject(s)
Hydrothermal Vents , Streptomyces , Anti-Bacterial Agents/chemistry , Streptomyces/chemistry , Hydrothermal Vents/microbiology , Antifungal Agents/pharmacology , Antifungal Agents/chemistry , Fungi , Molecular Structure
7.
BMC Infect Dis ; 23(1): 251, 2023 Apr 19.
Article in English | MEDLINE | ID: mdl-37076819

ABSTRACT

BACKGROUND: Pulmonary tuberculosis (TB) can impair pulmonary function (PF), especially in the elderly. The risk factors associated with the severity of PF impairment in the elderly with pulmonary TB remain unclear. Hence, this retrospective study aimed to address this issue to help improve the management of TB in the elderly population. METHODS: From January 2019 to February 2022, the elderly who were admitted to our hospital for pulmonary TB and underwent PF testing were included in this analysis. The forced expiratory volume in one second percent of predicted (FEV1% predicted) and clinical characteristics were collected and analyzed retrospectively. The extent of impaired PF was then categorized based on the FEV1% predicted and classified as grade 1-5. Logistic regression analysis was used to analyze the risk factors for impaired PF. RESULTS: A total of 249 patients who met the enrollment criteria were included in this analysis. According to the results of FEV1% predicted, all patients were classified as grade 1 (n = 37), grade 2 (n = 46), grade 3 (n = 55), grade 4 (n = 56), or grade 5 (n = 55). Statistical analysis showed that albumin (adjusted odds ratio (aOR) = 0.928, P = 0.013), body mass index (BMI) < 18.5 kg/m2 (aOR = 4.968, P = 0.046), lesion number ≥ 3 (aOR = 4.229, P < 0.001), male (aOR = 2.252, P = 0.009), respiratory disease (aOR = 1.669, P = 0.046), and cardiovascular disease (aOR = 2.489, P = 0.027) were related to the impairment of PF. CONCLUSIONS: PF impairment is common in the elderly with pulmonary TB. The male sex, BMI < 18.5 kg/m2, lesion number ≥ 3, hypoproteinemia, and respiratory and cardiovascular comorbidities were identified as risk factors for significant PF impairment. Our findings highlight the risk factors associated with PF impairment, which may be helpful to improve the current management of pulmonary TB in the elderly to save their lung function.


Subject(s)
Tuberculosis, Pulmonary , Tuberculosis , Humans , Male , Aged , Retrospective Studies , Lung , Tuberculosis/complications , Tuberculosis, Pulmonary/complications , Tuberculosis, Pulmonary/epidemiology , Forced Expiratory Volume , Risk Factors
8.
Fitoterapia ; 164: 105377, 2023 Jan.
Article in English | MEDLINE | ID: mdl-36544299

ABSTRACT

The ocean is a treasure house with rich resources of new chemical and biological molecules. A growing body of evidence suggests that marine fungi represent a huge and largely untapped resource of natural products that have been optimized by evolution for biological and ecological relevance. Alkaloids, the important components of natural products, have attracted much attention from medicinal and natural product chemists due to their unique structures and biological potential. The number and variety of alkaloids from marine fungi discovered in recent years maintain an upward trend. This review would give a systematic overview of the structures and bioactivities of marine fungal alkaloids obtained over the past six years and inspire the development of novel pharmaceutical agents.


Subject(s)
Alkaloids , Biological Products , Fungi/chemistry , Molecular Structure
9.
Phytochemistry ; 205: 113479, 2023 Jan.
Article in English | MEDLINE | ID: mdl-36270410

ABSTRACT

A previously unreported alkaloid, bearing an undescribed 5/7/8 tricyclic heterocyclic skeleton, shornephine D, an undescribed diketomorpholine (DKM) shornephine B, two undescribed diketomorpholine derivatives shornephine C and seco-shornephine B methyl ester, an undescribed indole-isoquinoline alkaloid asterresin C, three undescribed indole alkaloids asterresins A-B and D, together with five known compounds, were isolated from the culture of hydrothermal vent associated fungus Aspergillus terreus CXX-158-20. Their structures were unambiguously determined by nuclear magnetic resonance (NMR), mass spectrometry, Mosher's method, 13C NMR calculation in combination with DP4+, and ECD calculations. Shornephine D and asterresin C represent two undescribed heterocyclic skeletons. Asterresin D and giluterrin exhibited cytotoxicity activities with IC50 values of 3.96 µM and 7.97 µM against A549 cell line. Asterresin D exhibited cytotoxicity activities with IC50 values of 12.36 µM and 12.48 µM against Namalwa and U266 cell lines. Asterresin A and giluterrin exhibited synergistic effect with adriamycin against MCF-7 cell line.


Subject(s)
Alkaloids , Antineoplastic Agents , Hydrothermal Vents , Humans , Aspergillus/chemistry , MCF-7 Cells , Alkaloids/metabolism , Indole Alkaloids/metabolism , Antineoplastic Agents/pharmacology , Molecular Structure
10.
J Nat Prod ; 85(11): 2620-2625, 2022 11 25.
Article in English | MEDLINE | ID: mdl-36318598

ABSTRACT

We report the discovery of talaropeptins A (1) and B (2), tripeptides with an unusual 5/6/5 heterocyclic scaffold and an N-trans-cinnamoyl moiety, which were identified from the marine-derived fungus Talaromyces purpureogenus CX11. A bioinformatic analysis of the genome of T. purpureogenus CX11 and gene inactivation revealed that the biosynthesis of talaropeptins involves a nonribosomal peptide synthase gene cluster. Their chemical structures were elucidated using a combination of 1D and 2D NMR spectroscopy and mass spectrometry. The absolute configurations of 1 and 2 were established by electronic circular dichroism calculations and Marfey's method. The plausible biosynthesis of 1 and 2 is also proposed on the basis of gene deletion, substrate feeding, and heterologous expression. Compounds 1 and 2 showed moderate antifungal activity against phytopathogenic fungus Fusarium oxysporum with MIC values of 12.5 and 25 µg/mL, respectively.


Subject(s)
Antifungal Agents , Oligopeptides , Talaromyces , Antifungal Agents/chemistry , Antifungal Agents/isolation & purification , Antifungal Agents/pharmacology , Circular Dichroism , Magnetic Resonance Spectroscopy , Molecular Structure , Talaromyces/chemistry , Oligopeptides/chemistry , Oligopeptides/isolation & purification , Oligopeptides/pharmacology
11.
J Org Chem ; 87(19): 13270-13279, 2022 10 07.
Article in English | MEDLINE | ID: mdl-36131357

ABSTRACT

Five new unusual citrinin-derived alkaloids with a tetracyclic core, citrinidines A-E (1-5), two new amide alkaloids, methyl (2S,8E)-1'-(2-methyl-3-oxodec-8-enamido) butanoate (6) and (2S,8E)-2-methyl-3-oxodec-8-enamide (7), a new unusual citrinin trimer, tricitrinol C (8), a new citrinin acetal-ketal derivative, citrininol (9), together with four known citrinin monomers (10-13), and three known citrinin dimers (14-16), were isolated from the fermentation of hydrothermal vent-associated fungus Penicillium citrinum TW132-59. Their structures were unambiguously determined by nuclear magnetic resonance (NMR), mass spectrometry, Mosher's method, 13C NMR calculation in combination with DP4+, and ECD calculations. A plausible biosynthetic pathway of all new compounds (1-9) was proposed. Citrinin trimer (8) exhibited potent cytotoxicity activity with an IC50 value of 1.34 ± 0.11 µM, and compounds 1 and 15 showed moderate cytotoxicity with IC50 values of 17.50 ± 1.43 and 9.45 ± 0.55 µM, respectively, against A549 cell line.


Subject(s)
Alkaloids , Antineoplastic Agents , Citrinin , Hydrothermal Vents , Penicillium , Acetals , Alkaloids/chemistry , Alkaloids/pharmacology , Amides , Antineoplastic Agents/chemistry , Citrinin/chemistry , Citrinin/pharmacology , Fungi , Molecular Structure , Penicillium/chemistry
12.
PLoS One ; 17(9): e0274358, 2022.
Article in English | MEDLINE | ID: mdl-36136983

ABSTRACT

The dried plant material of medically important plant Actinidia eriantha especially when it remains in the form of powder often look morphologically similar to its related species. The lack of efficient methods to distinguish the authentic material from other similar species leads to chances of adulteration. The molecular authentication of herbal plant materials such as the internal transcribed spacer (ITS) sequences is considered as more reliable method compared to morphological traits. In this study, we aim to evaluate the potential of identification for roots of A. eriantha and its related species by ITS sequences. The lengths of ITS regions ranged from 624 to 636 bp with GC content ranging from 50.96% to 59.55%. A total of 194 variation sites and 46 haplotypes were formed in 185 samples. Among them, the roots of A. eriantha possessed specific sites at 85bp (C), 205bp (T), 493bp (C), 542bp (G), 574bp (C), 582bp (T) and 610bp (G), while A. hemsleyana, A. callosa, A. valvata and A. polygama have their own specific sites. The inter-specific genetic distance among 8 Actinidia species in the range 2.28% to 11.00%. The phylogenetic tree constructed with ITS, ITS1 and ITS2 region showed that the ITS sequences have higher potential for identification in 8 Actinidia species. However, as to A. eriantha, A. hemsleyana and A. valvata, these three barcodes have the same identification ability. The ITS regions indicated that different samples from same species can be grouped together, except for A. arguta and A. melanandrah. In conclusion, the ITS sequences can be used as an efficient DNA barcode for the identification of A. eriantha and its related species.


Subject(s)
Actinidia , Actinidia/genetics , Phylogeny , Plant Roots/genetics , Plants , Powders
13.
Eur J Med Chem ; 242: 114699, 2022 Nov 15.
Article in English | MEDLINE | ID: mdl-36001934

ABSTRACT

ß-Glucuronidase catalyzes the cleavage of glucuronosyl-O-bonds, whose inhibitors reduce the level of toxic substances present in the intestine caused by anti-cancer and anti-inflammatory therapies. Herein, we presented a new tool, Bioactive Fractions Filtering Platform (BFFP), which is able to reliably discern active candidate node from crude extracts. The source code for the BFFP is available on GitHub (https://github.com/BioGavin/msbff). With the assistant of BFFP, 25 gabosine and chlorogentisyl alcohol derivatives including 19 new compounds were isolated from a marine-derived fungus Epicoccum sp. GST-5. Compounds 7, 9-15 possessed an unusual hybrid skeleton of gabosine and chlorogentisyl alcohol units. Compounds 9-12, 16 and 17 possessed a novel three-membered spiral ring skeleton with one/two gabosine and one/two chlorogentisyl alcohol units. Compound 25 represented new gabosine-derived skeleton possessing an unusual 6/6/6/5/6 condensed ring system. All isolates were evaluated for in vitro E. coli ß-glucuronidase (EcGUS) inhibitory activity. 14 Compounds demonstrated superior inhibitory activity (IC50 = 0.24-4.61 µM) to that of standard d-saccharic acid 1,4-lactone (DSL, IC50 = 56.74 ± 4.01 µM). Compounds with chlorogentisyl alcohol moiety, such as 17 (IC50 = 0.24 ± 0.02 µM) and 1 (IC50 = 0.74 ± 0.03 µM), exhibited the most potent inhibitory activity. Furthermore, literature based QSAR profiling by applying PCA and OPLS analysis was carried out to analyze the features of compounds against EcGUS, revealing that the introduction of substituents able to form polar interactions with binding sites of receptor would lead to more active structures.


Subject(s)
Enzyme Inhibitors , Escherichia coli , Benzyl Alcohol , Complex Mixtures , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Fungi/metabolism , Glucaric Acid , Glucuronidase/metabolism , Informatics , Lactones , Molecular Docking Simulation , Molecular Structure , Structure-Activity Relationship
14.
Front Med (Lausanne) ; 9: 798465, 2022.
Article in English | MEDLINE | ID: mdl-35308541

ABSTRACT

Objective: To estimate the epidemic trends of tuberculosis (TB) in 30 high burden countries (HBCs) over the past 30 years, which is crucial for tracking the status of disease control, especially at the country level. Methods: Annual data on incidence and mortality of TB in these 30 HBCs were extracted from the Global Burden of Disease database. The average annual percent change (AAPC) was used to evaluate the trends of incidence and mortality. The trajectory analysis was used to identify different trends among the subgroup countries. The predicted incidence and mortality rates in 2025, 2030, and 2035 were also calculated. Results: The incidence and mortality decreased in most of the HBCs. The AAPCs of incidence ranged between -4.0 (Indonesia) and -0.2% (DR Congo) (all p < 0.05). The incidence trends in Lesotho (AAPC: 0%, 95% CI: -0.4, 0.3, p = 0.8) and South Africa (AAPC: -0.2%, 95% CI: -0.5, 0, p = 0.1) were stable, and increased in Kenya with AAPC of 0.1% (95% CI: 0.1, 0.2, p < 0.05). The AAPCs for mortality ranged between -5.8 (Ethiopia) and -0.6% (Central African Republic) (all p < 0.05). The mortality trends in DPR Korea (AAPC: 0.1%, 95% CI: -0.3, 0.4, p = 0.6) and Russian Federation (AAPC: -0.5%, 95% CI: -1.9, 0.9, p = 0.5) were stable, and increased in Lesotho and Zimbabwe with AAPC of 1.3% (95% CI: 1.1, 1.4, p < 0.05) and 1.6% (95% CI: 1.0, 2.2, p < 0.05), respectively. Trajectory analysis showed that the Central African Republic, Lesotho, Cambodia, Namibia, and South Africa had higher incidences, and the Central African Republic had higher mortality. Brazil and China had relatively lower rates of incidence and mortality. Predictions showed that reduction rates of incidence and mortality could hardly be reached compared with those set for the global targets for the majority HBCs. Conclusions: The disease burden of TB has been reduced among the majority HBCs over the last three decades. According to the current control levels, achieving the ambitious global targets at the country level for these 30 HBCs is challenging.

15.
Front Cell Infect Microbiol ; 11: 783568, 2021.
Article in English | MEDLINE | ID: mdl-35096640

ABSTRACT

BACKGROUND: Fever of unknown origin (FUO) is still a challenge for clinicians. Next-generation sequencing technologies, such as whole exome sequencing (WES), can be used to identify genetic defects in patients and assist in diagnosis. In this study, we investigated the application of WES in individuals with FUO. METHODS: We performed whole-exome sequencing on 15 FUO patients. Clinical information was extracted from the hospital information system. RESULTS: In 7/15 samples, we found positive results, including potentially causative mutations across eight different genes: CFTR, CD209, IRF2BP2, ADGRV 1, TYK2, MEFV, THBD and GATA2. CONCLUSIONS: Our results show that whole-exome sequencing can promote the genetic diagnosis and treatment of patients with FUO.


Subject(s)
Fever of Unknown Origin , Exome , Fever of Unknown Origin/genetics , High-Throughput Nucleotide Sequencing/methods , Humans , Mutation , Exome Sequencing/methods
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