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Objective: This study aims to estimate the overall in vitro activity of bedaquiline (BDQ) against clinical isolates of Mycobacterium abscessus complex (MABS) and M. avium complex (MAC), considering BDQ as a repurposed drug for non-tuberculous mycobacteria (NTM) infections. Methods: We conducted a systematic review of publications in PubMed/ MEDLINE, Web of Science, and Embase up to 15 April 2023. Studies were included if they followed the Clinical and Laboratory Standards Institute (CLSI) criteria for drug susceptibility testing (DST). Using a random effects model, we assessed the overall in vitro BDQ resistance rate in clinical isolates of MABS and MAC. Sources of heterogeneity were analysed using Cochran's Q and the I2 statistic. All analyses were performed using CMA V3.0. Results: A total of 24 publications (19 reports for MABS and 11 for MAC) were included. Using 1 µg/mL and 2 µg/mL as the breakpoint for BDQ resistance, the pooled rates of in vitro BDQ resistance in clinical isolates of MABS were found to be 1.8% (95% confidence interval [CI], 0.7–4.6%) and 1.7% (95% CI, 0.6–4.4%), respectively. In the case of MAC, the pooled rates were 1.7% (95% CI, 0.4–6.9%) and 1.6% (95% CI, 0.4–6.8%) for 1 µg/mL and 2 µg/mL, respectively. Conclusion: This study reports the prevalence of BDQ resistance in clinical isolates of MABS and MAC. The findings suggest that BDQ holds potential as a repurposed drug for treating MABS and MAC infections.

Abstract Interactions of tumor cells with immune cells in the tumor microenvironment play an important role during malignancy progression. We previously identified that GAS5 inhibited tumor development by suppressing proliferation of tumor cells in non-small cell lung cancer (NSCLC). Herein, we discovered a tumor-suppressing role for tumor cell-derived GAS5 in regulating tumor microenvironment. GAS5 positively coordinated with the infiltration of macrophages and T cells in NSCLC clinically, and overexpression of GAS5 promoted macrophages and T cells recruitment both in vitro and in vivo. Mechanistically, GAS5 stabilized p53 by directly binding to MYBBP1A and facilitating MYBBP1A-p53 interaction, and enhanced p53-mediated transcription of IRF1, which activated type I interferon signaling and increased the production of downstream CXCL10 and CCL5. We also found that activation of type I interferon signaling was associated with better immunotherapy efficacy in NSCLC. Furthermore, the stability of GAS5 was regulated by NAT10, the key enzyme responsible for N4-acetylcytidine (ac4C) modification, which bound to GAS5 and mediated its ac4C modification. Collectively, tumor cell-derived GAS5 could activate type I interferon signaling via the MYBBP1A-p53/IRF1 axis, promoting immune cell infiltration and potentially correlating with immunotherapy efficacy, which suppressed NSCLC progression. Our results suggested GAS5 as a promising predictive marker and potential therapeutic target for combination therapy in NSCLC. A schematic diagram demonstrating the regulatory effect of GAS5 on immune cell infiltration by activating type I interferon signaling via MYBBP1A-p53/IRF1 axis in non-small cell lung cancer. IFN, interferon.

To reduce the production costs of steel reinforced UHPC composite beams and fully utilize the material strength of steel, this paper introduced an innovative prefabricated composite beam comprising a reinforced ultra-high performance concrete beam and an internal castellated inverted T-steel, forming an inverted castellated T-steel reinforced UHPC (ICTSRU) composite beam. Investigating the shear behavior of ICTSRU beams is the primary objective of this research. Seven composite beams were loaded under simply supported four-point loading conditions, and the effects of key parameters such as the shear span-depth ratio, steel fiber volume fraction, and stirrup ratio were studied. The findings from the test indicated that the ICTSRU beams demonstrated superior deformation ability and significant residual shear strength after the peak load. The shear span-to-depth ratio significantly influenced the failure modes and carrying capacity of the ITSRU beams. As the span-to-depth ratio increased from 1.1 to 1.6 and 2.1, the failure mode developed from shear failure to flexural failure and the shear resistance decreased by 19 % and 48 %, respectively. Enhancing the steel fiber volume fraction and the stirrup ratio can mitigate the initiation and propagation of diagonal cracks, thereby augmenting the shear strength and ductility of ICTSRU beams. Following a comprehensive analysis of the experimental results, a predictive model for the shear strength of the ICTSRU beam is proposed. This model posits that the shear capacity of the ICTSRU beam comprises the shear resistance contributions from the UHPC matrix, stirrups, steel web, and steel fibers. A comparative analysis of the computed values and the test results showed that the proposed calculation method has a high accuracy in predicting the ultimate shear strength of ICTSRU beams.

Abstract Background Meta‐analyses have primarily focused on the effects of exercise‐based prehabilitation on postoperative outcomes and ignored the role of nutritional intervention. In this study, we filled this gap by investigating the effect of nutrition‐based prehabilitation on the postoperative outcomes of patients who underwent esophagectomy and gastrectomy. Methods Five electronic databases, namely, PubMed, the Web of Science, Embase, Cochrane Library, and CINAHL, were searched. Adults diagnosed with esophagogastric cancer who were scheduled to undergo surgery and had undergone uni‐ or multimodal prehabilitation, with at least a week of mandatory nutritional intervention, were included. Forest plots were used to extract and visualize the data from the included studies. The occurrence of any postoperative complication was considered the primary endpoint. Results Eight studies met the eligibility criteria, with five randomized controlled trials (RCTs) and three cohort studies. In total, 661 patients were included. Any prehabilitation, that is, unimodal (only nutrition) and multimodal prehabilitation, collectively decreased the risk of any postoperative complication by 23% (95% confidence interval [CI] = 0.66–0.90). A similar effect was exclusively observed for multimodal prehabilitation (risk ratio [RR] = 0.78, 95% CI = 0.66–0.93); however, it was not significant for unimodal prehabilitation. Any prehabilitation significantly decreased the length of hospital stay (LOS) (weighted mean difference = −0.77, 95% CI = −1.46 to −0.09). Conclusions Nutrition‐based prehabilitation, particularly multimodal prehabilitation, confers protective effects against postoperative complications after esophagectomy and gastrectomy. Our findings suggest that prehabilitation slightly decreases LOS; however, the finding is not clinically significant. Therefore, additional rigorous RCTs are warranted for further substantiation.

ABSTRACT This study aimed to assess the possible causes of discordant results between Xpert MTB/RIF (Xpert) and Bactec MGIT 960 Culture System (MGIT960) regarding rifampicin (RIF) susceptibility in Mycobacterium tuberculosis. Patients with previous RIF-resistant tuberculosis who were admitted to Wenzhou Central Hospital from January 2020 to December 2022 were enrolled. The isolates obtained from these patients were subjected to RIF susceptibility tests using Xpert and MGIT960, and the minimum inhibitory concentration (MIC) of RIF was determined by the MYCOTB MIC plate test. Additionally, molecular docking and molecular dynamics (MD) simulations were performed to evaluate the binding efficacy of rpoB and RIF based on rpoB mutations detected in the isolates with discordant RIF susceptibility results. A total of 28 isolates with discordant RIF susceptibility test results were detected, 15 of them were RIF susceptible with MICs ≤ 0.5 µg/mL. Twelve out of 15 isolates contained borderline RIF resistance-associated mutations [L430P (n = 6), H445N (n = 6)], 1 isolate had D435Y and Q429H double mutation, and the remaining 2 isolates had a silent (Q432Q) mutation. Compared with the affinity of RIF toward the wild type (WT) (−45.83 kcal/mol) by MD, its affinity toward L452P (−55.52 kcal/mol), D435Y (−47.39 kcal/mol), L430P (approximately −69.72 kcal/mol), H445N (−49.53 kcal/mol), and Q429H (−55.67 kcal/mol) increased. Borderline RIF resistance-associated mutations were the main cause for the discordant RIF susceptibility results between Xpert and MGIT960, and the mechanisms of the resistance need further investigated.IMPORTANCEThis study is aimed at assessing discordant results between Xpert MTB/RIF (Xpert) assay and Bactec MGIT 960 Culture System (MGIT960) regarding the detection of rifampicin (RIF)-resistant Mycobacterium tuberculosis isolates in Wenzhou, China. The discordant results of RIF between these two assays were mainly caused by borderline RIF resistance-associated mutations, subsequently by silent mutations of rpoB. Borderline RIF resistance- associated mutations detected in our study were demonstrated to not be affected by the affinity of rpoB and RIF by molecular dynamics, and the mechanism of resistance was needed to be clarified. For the discordant results of RIF by Xpert and MGIT960 that occurred, rpoB DNA sequencing was recommended to investigate its association with resistance to RIF.

Background: The prevalence of nontuberculous mycobacterial (NTM) disease in children is increasing worldwide. The clinical manifestations of pediatric NTM patients are significantly different from those of adult patients, but the knowledge of the disease is generally poor. Methods: English databases (PubMed, Web of Science, Embase, BIOSIS) and Chinese databases (CNKI, Wanfan, VIP) were searched on October 15th, 2022. All the articles of cross-sectional and cohort studies reporting the species composition and lesion site of the NTM disease in children using well-recognized NTM species identification methods were taken into account. Using a random effects model, we assessed the disease lesion sites and the prevalence of different NTM species in pediatric NTM disease. Sources of heterogeneity were analyzed using Cochran's Q and the I2 statistic. All analyses were performed using CMA V3.0. Results: The prevalence rates of NTM disease in children ranged between 0.6 and 5.36/100,000 in different countries, and Europe reported the highest prevalence rate. The most common clinical lesion site was lymph node, accounting for 71.1 % (55.0 %–83.2 %), followed by lung (19.3 %, 9.8%–34.4 %）and then skin and soft tissue (16.6 %,13.5%–20.3 %). Mycobacterium avium complex (MAC) was the most isolated NTM pathogen in children, accounting for 54.9 % (39.4%–69.6 %). Inconsistent with adult patients, Mycobacterium avium accounted for a dominant proportion in MAC than Mycobacterium intracellulare. Conclusions: The lymph node was the most affected organ in pediatric NTM disease, while Mycobacterium avium was the most isolated pathogenic species in children.

Sugarcane smut, a serious disease caused by the fungus Sporosorium scitamineum, can result in 30% to 100% cane loss. The most affordable and efficient measure of preventing and handling sugarcane smut disease is to select disease-resistant varieties. A comprehensive evaluation of disease resistance based on the incidence of smut disease is essential during the selection process, necessitating the rapid and accurate identification of sugarcane smut. Traditional identification methods, which rely on visual observation of symptoms, are time-consuming, costly, and inefficient. To address these limitations, we present the lightweight sugarcane smut detection model (YOLOv5s-ECCW), which incorporates several innovative features. Specifically, the EfficientNetV2 is incorporated into the YOLOv5 network to achieve model compression while maintaining high detection accuracy. The convolutional block attention mechanism (CBAM) is added to the backbone network to improve its feature extraction capability and suppress irrelevant information. The C3STR module is used to replace the C3 module, enhancing the ability to capture global large targets. The WIoU loss function is used in place of the CIoU one to improve the bounding box regression’s accuracy. The experimental results demonstrate that the YOLOv5s-ECCW model achieves a mean average precision (mAP) of 97.8% with only 4.9 G FLOPs and 3.25 M parameters. Compared with the original YOLOv5, our improvements include a 0.2% increase in mAP, a 54% reduction in parameters, and a 70.3% decrease in computational requirements. The proposed model outperforms YOLOv4, SSD, YOLOv5, and YOLOv8 in terms of accuracy, efficiency, and model size. The YOLOv5s-ECCW model meets the urgent need for the accurate real-time identification of sugarcane smut, supporting better disease management and selection of resistant varieties.

IntroductionEndophytes play a significant role in regulating plant root development and facilitating nutrient solubilization and transportation. This association could improve plant growth. The present study has uncovered a distinct phenotype, which we refer to as "white root", arising from the intricate interactions between endophytic fungi and bacteria with the roots in a sugarcane and bamboo fungus (Dictyophora indusiata) intercropping system.MethodsWe investigated the mechanisms underlying the formation of this “white root” phenotype and its impact on sugarcane yield and metabolism by metabarcoding and metabolome analysis.Results and DiscussionInitial analysis revealed that intercropping with D. indusiata increased sugarcane yield by enhancing the number of viable tillers compared with bagasse and no input control. Metabarcoding based on second-generation and third-generation sequencing indicated that D. indusiate and Bacillus aryabhattai dominates the fungal and bacterial composition in the “white root” phenotype of sugarcane root. The coexistence of D. indusiata and B. aryabhattai as endophytes induced plant growth-promoting metabolites in the sugarcane root system, such as lysoPC 18:1 and dihydrobenzofuran, probably contributing to increased sugarcane yield. Furthermore, the association also enhanced the metabolism of compounds, such as naringenin-7-O-glucoside (Prunin), naringenin-7-O-neohesperidoside (Naringin)*, hesperetin-7-O-neohesperidoside (Neohesperidin), epicatechin, and aromadendrin (Dihydrokaempferol), involved in flavonoid metabolism during the formation of the endophytic phenotype in the sugarcane root system. These observations suggest that the “white root” phenotype promotes sugarcane growth by activating flavonoid metabolism. This study reports an interesting phenomenon where D. indusiata, coordinate with the specific bacteria invade, forms a “white root” phenotype with sugarcane root. The study also provides new insights into using D. indusiata as a soil inoculant for promoting sugarcane growth and proposes a new approach for improve sugarcane cultivation.

Abstract Background Lodging seriously affects sugarcane stem growth and sugar accumulation, reduces sugarcane yield and sucrose content, and impedes mechanization. However, the molecular mechanisms underlying sugarcane lodging tolerance remain unclear. In this study, comprehensive transcriptomic and proteomic analyses were performed to explore the differential genetic regulatory mechanisms between upright (GT42) and lodged (GF98-296) sugarcane varieties. Results The stain test showed that GT42 had more lignin and vascular bundles in the stem than GF98-296. The gene expression analysis revealed that the genes that were differentially expressed between the two varieties were mainly involved in the phenylpropanoid pathway at the growth stage. The protein expression analysis indicated that the proteins that were differentially expressed between the two varieties were related to the synthesis of secondary metabolites, the process of endocytosis, and the formation of aminoacyl-tRNA. Time-series analysis revealed variations in differential gene expression patterns between the two varieties, whereas significant protein expression trends in the two varieties were largely consistent, except for one profile. The expression of CYP84A, 4CL, and CAD from the key phenylpropanoid biosynthetic pathway was enhanced in GT42 at stage 2 but suppressed in GF98-296 at the growth stage. Furthermore, the expression of SDT1 in the nicotinate and nicotinamide metabolism was enhanced in GT42 cells but suppressed in GF98-296 cells at the growth stage. Conclusion Our findings provide reference data for mining lodging tolerance-related genes that are expected to facilitate the selective breeding of sugarcane varieties with excellent lodging tolerance.

Abstract Tuberculosis (TB) is an airborne disease caused by Mycobacterium tuberculosis (Mtb). Whilst a functional role for humoral immunity in Mtb protection remains poorly defined, previous studies have suggested that antibodies can contribute towards host defense. Thus, identifying the critical components in the antibody repertoires from immune, chronically exposed, healthy individuals represents an approach for identifying new determinants for natural protection. In this study, we performed a thorough analysis of the IgG/IgA memory B cell repertoire from occupationally exposed, immune volunteers. We detail the identification and selection of a human monoclonal antibody that exhibits protective activity in vivo and show that it targets a virulence factor LpqH. Intriguingly, protection in both human ex vivo and murine challenge experiments was isotype dependent, with most robust protection being mediated via IgG2 and IgA. These data have important implications for our understanding of natural mucosal immunity for Mtb and highlight a new target for future vaccine development.

ABSTRACT: Objectives: Bedaquiline (BDQ) is a potent drug for treating drug-resistant tuberculosis (TB). Here, we analysed the resistance profiles of BDQ in CFZ-resistant clinical isolates and investigated the clinical risk factors of BDQ and CFZ cross/co-resistance. Methods: The AlarmarBlue microplate assay was performed to determine the minimum inhibitory concentration (MIC) of the CFZ-resistant Mycobacterium tuberculosis (MTB) clinical isolates to CFZ and BDQ. The clinical characteristics of the respective patients were analysed to explore the possible risk factors of BDQ resistance. The drug-resistance-associated genes including Rv0678, Rv1979c, atpE, pepQ and Rv1453 were sequenced and analysed. Results: A total of 72 clinical CFZ-resistant MTB isolates were collected; among these, half were identified as BDQ-resistant. The MIC value of BDQ closely correlated with CFZ (Spearman's q = 0.766, P < 0.005). Among the isolates with a MIC of CFZ ≥4 mg/L, 92.31% (12/13) were resistant to BDQ. Pre-XDR and exposure to BDQ or CFZ are the major risk factors for concurrent BDQ resistance. Among the 36 cross/co-resistant isolates, 50% (18/36) had mutations in Rv0678, 8.3% (3/36) had mutations in Rv0678+Rv1453, 5.6% (2/36) had mutations in Rv0678+Rv1979c, 2.8% (1/36) had mutations in Rv0678+Rv1979c+Rv1453, 2.8% (1/36) had mutations in atpE+Rv0678+Rv1453, 2.8% (1/36) had mutations in Rv1979c, and 27.7% (10/36) had no variations in the target genes. Conclusion: Nearly half of the CFZ-resistant isolates were still sensitive to BDQ, whereas this rate dramatically decreased among patients with pre-XDR TB or those who had been exposed to BDQ or CFZ.

ObjectivesMatrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a potent tool for detecting drug resistance in tuberculosis (TB); however, concerns about its reliability have been raised. In this study, we assessed the reliability of MassARRAY (Sequenom, Inc.), which is a MALDI-TOF MS-based method, by comparing it to the well-established GeneXpert assay (Cepheid) as a reference method.MethodsA retrospective study was conducted using laboratory data retrieved from Henan Chest Hospital (Zhengzhou, China). To ensure a rigorous evaluation, we adopted a comprehensive assessment approach by integrating multiple outcomes of the Xpert assay across various specimen types.ResultsAmong the 170 enrolled TB cases, MassARRAY demonstrated significantly higher sensitivity (85.88%, 146 of 170) compared to the Xpert assay (76.62%, 118 of 154) in TB diagnosis (p

Summary: Background: Human immunodeficiency virus (HIV) significantly increases the risk of non-Hodgkin lymphoma (NHL) development, yet the population-level impact on NHL burden is unquantified. We aim to quantify this association and estimate the global burden of HIV-associated NHL. Methods: In this meta-analysis, we searched five databases (PubMed, EMBASE, Cochrane Library, Web of Science, Scopus) from database inception up to September 13, 2023, identifying cohort, case–control, or cross-sectional studies with an effective control group to assess NHL risk among individuals with HIV infection, with two authors extracting summary data from reports. Global and regional HIV-associated population attributable fraction (PAF) and NHL disease burden were calculated based on the pooled risk ratio (RR). HIV prevalence and NHL incidence were obtained from the Joint United Nations Programme on HIV/AIDS (UNAIDS) and Global Burden of Diseases, Injuries, and Risk Factors Study 2019. Trends in NHL incidence due to HIV were assessed using age-standardised incidence rate (ASIR) and estimated annual percentage change (EAPC). This study was registered with PROSPERO (CRD42023404150). Findings: Out of 14,929 literature sources, 39 articles met our inclusion criteria. The risk of NHL was significantly increased in the population living with HIV (pooled RR 23.51, 95% CI 17.62–31.37; I2 = 100%, p

Introduction: Bone typing is crucial to enable the choice of a suitable implant, the surgical technique, and the evaluation of the clinical outcome. Currently, bone typing is assessed subjectively by the surgeon. Objective: The aim of this study is to establish an automatic quantification method to determine local bone types by the use of cone-beam computed tomography (CBCT) for an observer-independent approach. Methods: Six adult human cadaver skulls were used. The 4 generally used bone types in dental implantology and orthodontics were identified, and specific Hounsfield unit (HU) ranges (grey-scale values) were assigned to each bone type for identification by quantitative CBCT (qCBCT). The selected scanned planes were labelled by nonradiolucent markers for reidentification in the backup/cross-check evaluation methods. The selected planes were then physically removed as thick bone tissue sections for in vitro correlation measurements by qCBCT, quantitative micro–computed tomography (micro-CT), and quantitative histomorphometry. Results: Correlation analyses between the different bone tissue quantification methods to identify bone types based on numerical ranges of HU values revealed that the Pearson correlation coefficient of qCBCT with micro-CT and quantitative histomorphometry was R = 0.9 (P = .001) for all 4 bone types . Conclusions: We found that qCBCT can reproducibly and objectively assess human bone types at implant sites.

Objective: The resistance of Mycobacterium tuberculosis (Mtb) to currently available fluoroquinolones (FQs), namely ofloxacin (OFX), levofloxacin (LFX), and moxifloxacin (MFX), renders the treatment of TB infections less successful. In this study, we aimed to evaluate the susceptibility and intracellular killing assay of Mtb to next-generation FQs in vitro and determine the correlation of FQs resistance and newly detected mutations in gyrB by molecular docking. Methods: Antimicrobial susceptibility test was performed to determine the minimum inhibitory concentrations (MICs) of six FQs, including currently available FQs (OFX, LFX, and MFX) and next-generation FQs, i.e., sitafloxacin (SFX), finafloxacin (FIN) and delafloxacin (DFX) against Mtb clinical isolates obtained in 2015 and 2022, respectively. Quinolone-resistance-determining regions of gyrA and gyrB were subjected to DNA sequencing and the correlation of FQs resistance and new mutations in gyrB were determined by molecular docking. Furthermore, the intracellular antibacterial activity of the six FQs against Mtb H37Rv in THP-1 cells was evaluated. Results: SFX exhibited the highest antibacterial activity against Mtb isolates (MIC90 = 0.25 μg/mL), whereas DFX and OFX exhibited comparable activity (MIC90 = 8 μg/mL). A statistically significant difference was observed among the MICs of the new generation FQs (SFX, P = 0.002; DFX, P = 0.008). Additionally, a marked increase in MICs was found in strains isolated in 2022 compared with those isolated in 2015. There might be correlation between FQs resistance and mutations in gyrB G520T and G520A. Cross-resistance rate between SFX and MFX was 40.6 % (26/64). At a concentration of 1 μg/mL, SFX exhibited high intracellular antibacterial activity (96.6 % ± 1.5 %) against the Mtb H37Rv, comparable with that of MFX at a concentration of 2 μg/mL. Conclusion: SFX exhibits the highest inhibitory activity against Mtb in vitro and THP-1 cell lines, which exhibits partial-cross resistance with MFX. There might be correlation between FQs resistance and mutations in gyrB G520T and G520A.Our findings provide crucial insights into the potential clinical application of SFX and DFX in the treatment of Mtb infections.

Non-tuberculous mycobacteria (NTM) are opportunistic pathogens that can infect all body tissues and organs. In particular, the lungs are the most commonly involved organ, with NTM pulmonary diseases causing serious health issues in patients with underlying lung disease. Moreover, NTM infections have been steadily increasing worldwide in recent years. NTM are also naturally resistant to many antibiotics, specifically anti-tuberculosis (anti-TB) drugs. The lack of drugs targeting NTM infections and the increasing drug resistance of NTM have further made treating these mycobacterial diseases extremely difficult. The currently recommended NTM treatments rely on the extended indications of existing drugs, which underlines the difficulties of new antibiotic discovery against NTM. Another challenge is determining which drug combinations are most effective against NTM infection. To a certain extent, anti-NTM drug development depends on using already available antibiotics and compounds. Here, we aimed to review new antibiotics or compounds with good antibacterial activity against NTM, focusing on their mechanisms of action, in vitro and in vivo antibacterial activities.

Objectives: Xpert Mycobacterium tuberculosis/rifampin (MTB/RIF) Ultra (Xpert-Ultra) has shown better sensitivity in comparison with Xpert MTB/RIF (Xpert) in extrapulmonary tuberculosis (TB), whereas the head-to-head comparison of these methods in TB lymphadenitis had barely been performed. Methods: Patients with undiagnosed lymphadenopathy were recruited prospectively and consecutively, and fine-needle aspiration (FNA) biopsy or lymph node tissue was collected. The specimen was subjected to smear, culture, Xpert, and Xpert-Ultra assays. Culture and/or smear for acid-fast bacilli (AFB) or AFB observed on histopathology were performed as a reference. Results: A total of 106 participants were recruited, including 41 confirmed TB, 33 probable TB, and 32 non-TB lymphadenopathies. The head-to-head comparison for MTB detection showed that Xpert-Ultra produced the highest sensitivity when compared with smear, culture, and Xpert (75.7% vs 5.4 %, 13.5%, and 48.7%). When Xpert-Ultra outcomes were integrated for diagnosis, the percentage of confirmed TB lymphadenitis cases increased from 55.4% (41/74) to 85.1% (63/74). The sensitivities of Xpert-Ultra and Xpert on tissue were 73.6% (95% CI: 59.4-84.3) and 39.6% (95% CI: 26.8-54.0), respectively. The sensitivity of Xpert-Ultra on FNA samples (81.0%, 95% CI: 57.4-93.7) was higher than that of Xpert (71.4%, 95% CI: 47.7-87.8). Conclusion: Xpert-Ultra detected significantly more TB lymphadenitis cases than Xpert or culture. This superiority was particularly distinct using lymph node tissue than FNA detection.

Since the onset of the COVID-19 pandemic in 2020, global efforts towards tuberculosis (TB) control have encountered unprecedented challenges. There is an urgent demand for efficient and cost-effective diagnostic technologies for TB. Recent advancements in CRISPR–Cas technologies have improved our capacity to detect pathogens. The present study established a CRISPR–Cas12a-based multiplex detection (designated as MCMD) that simultaneously targets two conserved insertion sequences (IS6110 and IS1081) to detect Mycobacterium tuberculosis complex (MTBC). The MCMD integrated a graphene oxide-assisted multiplex recombinase polymerase amplification (RPA) assay with a Cas12a-based trans-cleavage assay identified with fluorescent or lateral flow biosensor (LFB). The process can be performed at a constant temperature of around 37°C and completed within 1 h. The limit of detection (LoD) was 4 copies μL−1, and no cross-reaction was observed with non-MTBC bacteria strains. This MCMD showed 74.8% sensitivity and 100% specificity in clinical samples from 107 patients with pulmonary TB and 40 non-TB patients compared to Xpert MTB/RIF assay (63.6%, 100%). In this study, we have developed a straightforward, rapid, highly sensitive, specific, and cost-effective assay for the multiplex detection of MTBC. Our assay showed superior diagnostic performance when compared to the widely used Xpert assay. The novel approach employed in this study makes a substantial contribution to the detection of strains with low or no copies of IS6110 and facilitates point-of-care (POC) testing for MTBC in resource-limited countries.

ABSTRACT This study evaluated the diagnostic performance of stool-based Xpert MTB/RIF Ultra assay (Xpert-Ultra, Cepheid, USA) against other tests using respiratory tract specimens (RTS) and stool for diagnosing adult pulmonary tuberculosis. A prospective study on patients with presumptive pulmonary tuberculosis was conducted in Beijing Chest Hospital from June to November 2021. The smear test, MGIT960 liquid culture, and Xpert MTB/RIF (Xpert, Cepheid, USA) were performed simultaneously on RTS, and smear, culture Xpert, and Xpert-Ultra were performed simultaneously using stool. Patients were grouped based on the outcomes of RTS examination and other tests. In total, 130 eligible patients were enrolled that included 96 pulmonary tuberculosis and 34 non-TB patients. The sensitivity of smear, culture, Xpert, and Xpert-Ultra using stool was 10.96%, 23.28%, 60.27%, and 79.45%, respectively. The specificities of Xpert and Xpert-Ultra using RTS and stool were all 100% (34/34). Notably, all five confirmed cases detected by bronchoalveolar lavage fluid (BALF) examination yielded Xpert-Ultra positive outcomes with the stool specimens. Xpert-Ultra assay on stool sample harbors comparable sensitivity with Xpert on RTS. Thus, the Xpert-Ultra testing on stool specimens could be a very promising and practical strategy to improve pulmonary tuberculosis (PTB) diagnosis, especially among patients who could not expectorate sputum. IMPORTANCE This study is aimed at assessing the value of Xpert MTB/RIF Ultra (Xpert-Ultra) in PTB on stool in adult in low HIV settings and Xpert-Ultra assay on stool sample harboring comparable sensitivity with Xpert MTB/RIF on respiratory tract specimens. Although the yield in stool samples by Xpert-Ultra is lower than RTS, it may be useful in detecting disease in presumptive TB patients who cannot expectorate sputum and are not open to BALF collection. In addition, Xpert-Ultra with a “trace call” on stool in adult was highly supportive of PTB.

To ensure the serviceable popularization of RHA and avoid the alkali silica reaction of mortar blended RHA, the impact of RHA particle size and dosage on the mechanical strength and alkali silica reaction expansion of the mortar prepared with nonreactive silica sand and river sand has been evolutionarily investigated. The hydrates products of the mortar containing RHA were conducted by thermogravimetric (TG) and X-ray diffraction analysis (XRD). The microstructure of the mortar containing RHA was also explored by scanning electron microscope with an energy-dispersive X-ray spectroscopy analysis (SEM-EDX). The experiment results demonstrate that the compressive strength of mortar could be maximum enhanced by 21.1 % when RHA8.8 substituted 20 % of cement due to the pozzolanic effect and physical filling effect of RHA. Moreover, RHA with massive amorphous silica could react with Na ions in pore solution to form alkali-silica gel, thereafter resulting in expanding and destroying the microstructure of the mortar. The real ASR expansion value fabricated by RHA increased as the RHA particle size increased, which was mostly up to 0.0177 % when 20 % cement was replaced by RHA with a mean particle of 27.6 µm. Cracks noticeably emerged on the surface and edge of the mortar containing RHA with the particle size from 8.8 µm to 27.6 µm when RHA substituted 20 % of cement (by mass). Overall, the RHA particle size should be lower than 8.8 µm, while the substitution of cement by RHA also be below 10 % of cement, which can safely avoid the destructive expansion of RHA in an alkali environment.

IntroductionGreater amounts of fertilizer are applied every year to meet the growing demand for food. Sugarcane is one of the important food sources for human beings.MethodsHere, we evaluated the effects of a sugarcane—Dictyophora indusiata (DI) intercropping system on soil health by conducting an experiment with three different treatments: (1) bagasse application (BAS process), (2) bagasse + DI (DIS process), and (3) the control (CK). We then analyzed soil chemistry, the diversity of soil bacteria and fungi, and the composition of metabolites to clarify the mechanism underlying the effects of this intercropping system on soil properties.Results and discussionSoil chemistry analyses revealed that the content of several soil nutrients such as nitrogen (N) and phosphorus (P) was higher in the BAS process than in the CK. In the DIS process, a large amount of soil P was consumed by DI. At the same time, the urease activity was inhibited, thus slowing down the loss of soil in the DI process, while the activity of other enzymes such as β-glucosidase and laccase was increased. It was also noticed that the content of lanthanum and calcium was higher in the BAS process than in the other treatments, and DI did not significantly alter the concentrations of these soil metal ions. Bacterial diversity was higher in the BAS process than in the other treatments, and fungal diversity was lower in the DIS process than in the other treatments. The soil metabolome analysis revealed that the abundance of carbohydrate metabolites was significantly lower in the BAS process than in the CK and the DIS process. The abundance of D(+)-talose was correlated with the content of soil nutrients. Path analysis revealed that the content of soil nutrients in the DIS process was mainly affected by fungi, bacteria, the soil metabolome, and soil enzyme activity. Our findings indicate that the sugarcane–DIS intercropping system can enhance soil health.

Abstract Background Miliary tuberculosis (TB) is one of the severest manifestations of TB that can be lethal when concomitant with the central nervous system (CNS) involvement. Bacteriological, biochemical and radiological methods for find CNS comorbidity in miliary TB was evaluated in this study. Methods Consecutive miliary TB adults were retrospectively enrolled from two designated TB hospitals in China. The capacities of examinations of cerebrospinal fluid (CSF), cerebral computed tomography (CT) and magnetic resonance imaging (MRI) for diagnosis of CNS involvement were assessed. Results Assessment of CNS involvement with a lumbar puncture and/or neuroimaging was undertaken in 282 out of 392 of acute miliary TB. Of these 282 patients, 87.59% (247/282) had CNS involvement. Cerebral contrast-enhanced MRI (96.05%, 170/177) and MRI (93.15%, 204/219) yielded significantly higher sensitivities over CSF examination (71.92%, 146/203, P

ABSTRACT Infections caused by rapidly growing mycobacteria (RGM) have increased globally. Chemotherapy against these infections is challenging due to the minimal antimicrobial choices available. The main aim of this study was to evaluate the in vitro susceptibilities of four tetracyclines against different RGM species. The MICs of eravacycline (ERC), omadacycline (OMC), sarecycline (SAC), and tigecycline (TGC) against the reference strains of 27 RGM species and 121 RGM clinical isolates were determined by microtiter plate assay. The minimum bactericidal concentrations (MBCs) and cytotoxicities of these antibiotics were also tested. Except for SAC, the other three tetracyclines had MICs of ≤0.5 μg/mL against all 27 RGM reference strains. ERC generally presented the lowest MICs, with MIC90s against the clinical isolates of Mycobacterium abscessus subsp. abscessus, Mycobacterium abscessus subsp. massiliense, and Mycobacterium fortuitum of 0.25 μg/mL, 0.25 μg/mL, and 0.06 μg/mL, respectively. TGC and OMC also showed equivalent in vitro inhibitory activities against the isolates, while the TGC MIC90s for M. abscessus subsp. abscessus, M. abscessus subsp. massiliense, and M. fortuitum were lower than or equal to the OMC MIC90s (1, 1, and 0.25 μg/mL versus 1, 2, and 2 μg/mL). In addition, the MIC50s of three of the antibiotics for each species were always 2-fold lower than the corresponding MIC90s. MBC and cytotoxicity assays indicated that all four tetracycline antibiotics tested were bacteriostatic agents with low toxicity to the THP-1 cell line. Tetracycline antibiotics are efficacious in RGM infection treatment, with omadacycline showing the best promise for clinical application due to its potent antimicrobial activity, safety, and convenient administration route. IMPORTANCE The global rise in antibiotic-resistant nontuberculous mycobacteria has prompted the urgent need for new antimicrobials, especially oral antibiotics. Currently, adverse effects have limited the use of tetracycline-class antibiotics, particularly tigecycline (TGC), in the treatment of rapidly growing mycobacteria (RGM). However, several new tetracycline-class antibiotics might overcome the limitations of TGC. We assessed the in vitro antibiotic susceptibilities of four tetracyclines (eravacycline, omadacycline, sarecycline, and tigecycline) against reference RGM strains and clinical isolates of different RGM species. We showed that three of these antibiotics (tigecycline, eravacycline, and omadacycline) might be efficacious in M. abscessus subsp. abscessus, M. abscessus subsp. massiliense, and M. fortuitum treatment. Furthermore, omadacycline was more promising for clinical application for M. abscessus infections as an oral drug, whereas sarecycline, which had the best safety parameters, should be considered a potential antibiotic for M. abscessus infections caused by susceptible strains. Our work underscores the possible clinical applications of tetracycline-class antibiotics in the treatment of RGM infections.

IntroductionThe objective of the study was to identify the causes of smear-positive-culture-negative (S+/C–) outcomes of patients with tuberculosis during the treatment course.MethodsA laboratory-based retrospective study was performed at the Beijing Chest Hospital in China. Within the study period, all patients with pulmonary tuberculosis (PTB) who undertook anti-TB treatments and yielded smear positive outcomes with simultaneous culture outcomes on sputa were considered. Patients were classified into three groups: (I) performed LJ medium culture only; (II) performed BACTEC MGIT960 liquid culture only; and (III) performed both LJ culture and MGIT960 culture. The S+/C– rates of each group were analyzed. The clinical medical records regarding patient category, follow-up bacteriologic examination data, and treatment response were investigated.ResultsIn total, 1,200 eligible patients were enrolled, and the overall S+/C– rate was 17.5% (210/1,200). Group I had obviously higher S+/C– rate (37%) than group II (18.5%) and group III (9.5%). When solid and liquid cultures were considered independently, the S+/C– outcome was observed more frequently in the solid culture group than in the liquid culture group (30.4%, 345/1,135 vs. 11.5%, 100/873; p < 0.001, χ2 = 102.64). Among the 102 S+/C– patients who had follow-up cultures performed, 35 (34.3%) had positive culture outcomes. Whereas among the 67 patients with follow-up information for more than 3 months but without supportive bacteriological evidence, 45 (67.2%, 45/67) had unfavorable prognosis (including relapse and unimproved conditions), and only 22 (32.8%, 22/67) patients had improved conditions. Compared with new cases, retreated cases produced S+/C– outcomes more frequently and had more chances to be cultivated bacilli successfully afterward.ConclusionsAmong our patients, the sporadic smear positive and culture negative outcomes for sputa are more likely associated with the technical failures of culture than with dead bacilli, and this is especially noteworthy for LJ medium culture.

ABSTRACT Multidrug-resistant tuberculosis (MDR-TB) is often associated with poor clinical outcomes. In this study, we evaluated the potential of nosiheptide (NOS) as a new drug candidate for treating Mycobacterium tuberculosis infections, including MDR-TB. The antimicrobial susceptibility testing was performed to determine the MICs of NOS against 18 reference strains of slowly growing mycobacteria (SGM) and 128 clinical isolates of M. tuberculosis. The postantibiotic effects (PAE) and interaction with other antituberculosis drugs of NOS were also evaluated using M. tuberculosis H37Rv. Fifteen out of the 18 tested reference strains of SGM had MICs far below 1 μg/mL. From the 128 M. tuberculosis clinical isolates, the MIC50 and MIC90 were 0.25 μg/mL and 1 μg/mL, respectively; the tentative epidemiological cutoff (ECOFF) was defined at 1 μg/mL. Furthermore, a Lys89Thr mutation was found in one M. tuberculosis isolate with a MIC of NOS >8 μg/mL. After 24 h of incubation, NOS at 1 μg/mL inhibited 25.79 ± 1.22% of intracellular bacterial growth, which was comparable with the inhibitory rate of 25.71 ± 3.67% achieved by rifampin at 2 μg/mL. Compared to rifampicin and isoniazid (INH), NOS had a much longer PAE, i.e., a value of about 16 days. In addition, a partial synergy between NOS and INH was observed. NOS has potent inhibitory activities against M. tuberculosis in vitro as well as in macrophages. Furthermore, the long PAE and partial synergistic effect with INH, in addition to the added safety of long-term use as a feed additive in husbandry, provide support for NOS being a promising drug candidate for tuberculosis treatment. IMPORTANCE This study is aimed at chemotherapy for MDR-TB, mainly to explore the anti-TB activity of the existing chemotherapeutic reagent. We found that NOS has potent inhibitory activities against M. tuberculosis in vitro regardless of the drug-resistant profile. Furthermore, NOS also showed the long PAE and partial synergistic effect with INH and is nontoxic, providing support for its promise as a drug candidate for drug-resistant tuberculosis treatment.

ABSTRACT Sudapyridine (WX-081) is a structural analog of bedaquiline (BDQ), which shows an anti-tuberculosis activity but, unlike BDQ, did not prolong QT interval (QT) in animal model studies. This study evaluated the antimicrobial activity of this novel drug against non-tuberculous mycobacteria (NTM). Fifty reference strains of different mycobacterial species, and 132 NTM clinical isolates from four commonly isolated NTM species were recruited. The microplate alamarBlue assay was performed to determine the MIC of WX-081 and BDQ. Cytotoxicity assay was performed for both drugs using the THP-1 cells, and the minimum bactericidal concentrations (MBCs) of both drugs against the reference strains of five selected NTM species were also determined. All the tested reference strains had MICs lower than 0.5 μg/mL, with the majority having MICs far below 0.1 μg/mL for WX-081. The epidemiological cut-offs of WX-081 ranged from 0.0156 μg/mL to 0.25 μg/mL against commonly isolated NTM, and this value was comparable with that of BDQ. The MBC/MIC ratios suggest a bacteriostatic activity for both drugs against the five selected NTM species. Cytotoxicity assays indicated that THP-1 cells had nearly 100% viability when exposed to WX-081 for 24 h below 4 μg/mL, 200- to 300-fold the MICs of Mycobacterium intracellulare, Mycobacterium avium, and Mycobacterium kansasii. WX-081 has a strong antimicrobial activity against different NTM species with low cytotoxicity and therefore has the potential to be used for treating NTM infections. IMPORTANCE Due to the rapidly increased cases globally, non-tuberculous mycobacteria (NTM) disease has become a significant public health problem. Over 200 species or subspecies of NTM have been reported, whereas pulmonary diseases in humans are caused mainly by M. avium complex (MAC), M. kansasii, and M. abscessus. Treatment of NTM infection is often challenging as natural resistance to most antibiotics is quite common among different NTM species. Hence, identifying highly active anti-NTM agents is a priority for potent regimen establishment. The pursuit of new drugs to treat multidrug-resistant-tuberculosis (MDR-TB) may also identify some agents with strong activity against NTM. Sudapyridine (WX-081) is a structural analog of bedaquiline (BDQ), which was developed to retain the antituberculosis efficacy but eliminate the severe side effect of BDQ. This study initially evaluated the antimicrobial activity of this novel drug against non-tuberculous mycobacteria (NTM).

ObjectiveTo investigate the in vitro activities of five oxazolidinones in parallel against the reference strains of different mycobacterial species and clinical isolates of Mycobacterium tuberculosis (Mtb), and shed light on the differences in the efficacy of these homolog drugs.Materials and methodsThe minimum inhibitory concentrations (MICs) of linezolid, tedizolid, sutezolid, delpazolid, and contezolid against 16 mycobacterial reference strains and 69 M. tuberculosis clinical isolates, including 17 drug-susceptible isolates and 52 multidrug-resistant (MDR) isolates, were determined by microplate alamarBlue assay (MABA). The intracellular killing activities of contezolid and linezolid against Mtb H37Rv were compared. In addition, mutations in the linezolid resistance-related genes (rplC, rplD, and 23S rRNA) of the Mtb clinical isolates were also analyzed.ResultsTedizolid exhibited the strongest inhibitory activities against the reference strains of both rapidly growing mycobacteria (RGM) and slowly growing mycobacteria (SGM), among the tested oxazolidinones. In contrast, sutezolid only manifested potent activity against reference strains of SGM. Linezolid, delpazolid, and contezolid were less active against the non-tuberculous mycobacterial references. For the Mtb clinical isolates, the antimicrobial action was ranked as: sutezolid > tedizolid > contezolid and linezolid > delpazolid, whereas no difference between drug-sensitive and multiple drug-resistant isolates was observed. Notably, contezolid demonstrated obviously superior intracellular antimicrobial activity than linezolid. Few strains harbored mutations in rrl gene or rplD genes, although these strains had drug susceptible profiles to linezolid.ConclusionDifferent oxazolidinones can have discrepant antimicrobial activity against different mycobacterial species, or have different manifestations out of cell or in cell. Understanding these differences would be helpful in choosing the appropriate drug in clinical practice.

While embedding in the global value chain (GVC), avoiding becoming a “pollution shelter” is an urgent issue that needs to be solved for the low-carbon economic development of countries along the Belt and Road. This study empirically analyzes the impact of manufacturing GVC embedding on carbon emissions and its mechanism from the perspectives of the embedding position and degree of the GVC. It then carries out group regressions by different industries, value chain embedding modes, and so on. Finally, this study constructs static and dynamic panel models to verify the nonlinear relationship between manufacturing GVC embedding and carbon emissions. The results show that the improvement of the embedding position of GVC in the manufacturing industry promotes an increase in carbon emissions; the deeper the embedding degree of GVC, the more conducive it is to carbon emissions reduction. Compared with the service industry, the impact of manufacturing GVC embedding on carbon emissions is more significant; regardless of the value chain embedding model, the deeper the embedding degree, the more conducive it is to carbon emissions reduction. Technological progress can weaken the impact of GVC embedding on carbon emissions. There is a U-shaped relationship between the GVC embedding position and carbon emissions in the manufacturing industry and an inverted U-shaped relationship between the GVC embedding degree and carbon emissions. This study provides beneficial insights for countries along the Belt and Road to embed into the GVC to avoid falling into the dilemma of “pollution growth”.

In recent years, with the rapid development of technology, artificial intelligence (AI) security issues represented by adversarial sample attack have aroused widespread concern in society. Adversarial samples are often generated by surrogate models and then transfer to attack the target model, and most AI models in real-world scenarios belong to black boxes; thus, transferability becomes a key factor to measure the quality of adversarial samples. The traditional method relies on the decision boundary of the classifier and takes the boundary crossing as the only judgment metric without considering the probability distribution of the sample itself, which results in an irregular way of adding perturbations to the adversarial sample, an unclear path of generation, and a lack of transferability and interpretability. In the probabilistic generative model, after learning the probability distribution of the samples, a random term can be added to the sampling to gradually transform the noise into a new independent and identically distributed sample. Inspired by this idea, we believe that by removing the random term, the adversarial sample generation process can be regarded as the static sampling of the probabilistic generative model, which guides the adversarial samples out of the original probability distribution and into the target probability distribution and helps to boost transferability and interpretability. Therefore, we proposed a score-matching-based attack (SMBA) method to perform adversarial sample attacks by manipulating the probability distribution of the samples, which showed good transferability in the face of different datasets and models and provided reasonable explanations from the perspective of mathematical theory and feature space. Compared with the current best methods based on the decision boundary of the classifier, our method increased the attack success rate by 51.36% and 30.54% to the maximum extent in non-targeted and targeted attack scenarios, respectively. In conclusion, our research established a bridge between probabilistic generative models and adversarial samples, provided a new entry angle for the study of adversarial samples, and brought new thinking to AI security.

Background: The natural resistance of rapidly growing mycobacteria (RGM) to multiple antibiotics renders the treatment of the infections caused less successful. The objective of this study was to evaluate the in vitro susceptibilities of four oxazolidinones against different RGM species. Methods: The microplate alamarBlue assay was performed to identify the minimum inhibitory concentrations (MICs) of four oxazolidinones – delpazolid, sutezolid, tedizolid, and linezolid – for 32 reference strains and 115 clinical strains of different RGM species. The MIC breakpoint concentration was defined as 16 μg/ml for linezolid. Next, the gene fragments associated with oxazolidinone resistance were amplified and sequenced, and mutations were defined in contrast with the sequences of the reference strains. Results: Tedizolid showed the strongest inhibitory activity against the Mycobacterium abscessus isolates. Delpazolid exhibited better antimicrobial activity against the Mycobacterium fortuitum isolates when compared to linezolid, with 4-fold lower MIC values. The protein alignment and structure-based analysis showed that there might be no correlation between oxazolidinone resistance and mutations in the rplC, rplD, and 23S rRNA genes in the tested RGM. Conclusions: Tedizolid had the strongest inhibitory activity against M. abscessus in vitro, while delpazolid presented the best inhibitory activity against M. fortuitum. This provides important insights into the potential clinical application of oxazolidinones to treat RGM infections.

Abstract Metastasis suggests a poor prognosis for cancer patients, and treatment strategies for metastatic cancer are still very limited. Numerous studies have shown that cancer-associated fibroblasts (CAFs), a large component of the tumor microenvironment, contribute to tumor metastasis. Stromal fibroblasts at metastatic sites are different from CAFs within primary tumors and can be termed metastasis-associated fibroblasts (MAFs), and they also make great contributions to the establishment of metastatic lesions and the therapeutic resistance of metastatic tumors. MAFs are capable of remodeling the extracellular matrix of metastatic tumors, modulating immune cells in the tumor microenvironment, promoting angiogenesis and enhancing malignant tumor phenotypes. Thus, MAFs can help establish premetastatic niches and mediate resistance to therapeutic strategies, including immunotherapy and antiangiogenic therapy. The results of preclinical studies suggest that targeting MAFs can alleviate the progression of metastatic cancer and mitigate therapeutic resistance, indicating that MAFs are a promising target for metastatic cancer. Here, we comprehensively summarize the existing evidence on MAFs and discuss their origins, generation, functions and related therapeutic strategies in an effort to provide a better understanding of MAFs and offer treatment perspectives for metastatic cancer.

ABSTRACT Due to the probability of decreased specificity, the practical value of performing the Xpert MTB/RIF Ultra (Xpert Ultra) assay over the Xpert assay for diagnosing pulmonary tuberculosis (TB) and rifampicin (RIF) resistance in a high TB burden setting was evaluated. Participants were recruited consecutively in three tertiary hospitals in China and allocated to the TB case detection and/or rifampicin (RIF) resistance detection group. Each sputum specimen was subjected to smear, MGIT960 liquid culture, and Xpert, and Xpert Ultra assay in parallel. Drug susceptibility testing was conducted for all recovered isolates in the RIF resistance detection group. In total, 1,079 patients were recruited to the case detection group and 450 to the RIF resistance detection group. Xpert Ultra had higher sensitivity than Xpert (92.26%, 322/349 versus 89.40%, 312/349; P = 0.006), whereas the most prominent increase was identified in the smear-negative patients (83.70% versus 78.52%; P = 0.039). The specificity of Xpert Ultra was slightly lower than that of Xpert (96.30%, 495/514 versus 98.25%, 505/514; P = 0.055). Reclassifying trace results as negative resulted in a 4.01% loss of sensitivity (from 92.26% to 88.25%) accompanied by a 1.37% gain in specificity (from 96.30% to 97.67%). Both the sensitivity (97.64% versus 99.21%, P = 0.313) and specificity (96.90% versus 97.21%, P = 0.816) of Xpert Ultra and Xpert for detection RIF resistance were comparable. In conclusion, Xpert Ultra could improve the diagnosis of smear-negative pulmonary TB in contrast to the Xpert assay. A high percentage of TB history did not significantly decrease the specificity of the test, which supports the potential role of Xpert Ultra as an initial diagnostic tool for TB. IMPORTANCE Xpert Ultra is more sensitive than Xpert, especially in smear-negative TB. A high percentage of TB history in the non-TB population did not significantly affect the reliability of the assay, which supports the potential role of Xpert Ultra as an initial diagnostic tool for TB.

The high level of inherent drug resistance of Mycobacterium abscessus makes the infection caused by it very difficult to be treated. The objective of this study was to evaluate the potential of nosiheptide (NOS) as a new drug candidate for treating M. abscessus infections. The microplate AlamarBlue assay was performed to determine the minimum inhibitory concentrations (MICs) of NOS for 28 reference strains of rapidly growing mycobacteria (RGM) and 77 clinical isolates of M. abscessus. Time-kill kinetic and post-antibiotic effect (PAE) of NOS against M. abscessus was evaluated. Its bactericidal activity against M. abscessus in macrophages was determined by an intracellular colony numerating assay. NOS manifested good activity against the reference strains of RGM and M. abscessus clinical isolates in vitro. The MICs of NOS against M. abscessus clinical isolates ranged from 0.0078 to 1 μg/ml, and the MIC50 and MIC90 were 0.125 μg/ml and 0.25 μg/ml, respectively. The pattern of growth and kill by NOS against M. abscessus was moderate with apparent concentration-dependent characteristics, and the PAE value of NOS was found to be ~6 h. Furthermore, NOS had low cell toxicity against the THP-1 cell line after 48 h of exposure (IC50 = 106.9 μM). At 4 μg/ml, NOS exhibited high intracellular bactericidal activity against M. abscessus reference strains with an inhibitory rate of 66.52% ± 1.51%, comparable with that of clarithromycin at 2 μg/ml. NOS showed suitable inhibitory activities against M. abscessus in vitro and in macrophages and could be a potential drug candidate to treat M. abscessus infection.

BackgroundPulmonary non-tuberculous mycobacteria (NTM) infection has become a public health concern in China and around the world. The objective of this study was to describe the longitudinal changes in the frequency and diversity of NTM in northern China.MethodsWe retrospectively analyzed data on mycobacterium species in Beijing Chest Hospital from January 2014 to December 2021. The isolates were identified to species level by targeted DNA sequencing.ResultsAfter excluding duplicates, 1,755 NTM strains were analyzed, which were from 27 provinces in China over 8 years. Among all mycobacteria, the proportion of NTM increased each year, from 4.24% in 2014 to 12.68% in 2021. Overall, 39 different NTM species were identified, including 23 slow growing mycobacteria (SGM) and 16 rapid growing mycobacteria (RGM). The most common species were M. intracellulare (51.62%), M. abscessus (22.22%), M. kansasii (8.32%), M. avium (7.75%) and M. fortuitum (2.05%). The number of NTM species identified also increased each year from 9 in 2014 to 26 in 2021. Most species showed stable isolation rates over the years; however, the proportion of M. avium increased from 3.85 to 10.42% during the study period. Besides, 81 non-mycobacteria strains, including Gordonia (21 isolates), Nocardia (19 isolates) and Tsukamurella (17 isolates), etc., were also discovered.ConclusionThe proportion of NTM and species diversity increased considerably in northern China from 2014 to 2021. M. intracellulare was the most common NTM isolated among respiratory specimens, followed by M. abscessus and M. kansasii. Rare NTM species and non-mycobacteria pathogens also need attention.

Introduction: To evaluate the performance of Xpert MTB/RIF Ultra (Xpert-Ultra) in testing pleural tissue and fluid collected by medical thoracoscopy among patients with unexplained exudative pleural effusion. Methods: Patients with an undiagnosed exudative pleural effusion were prospectively and consecutively recruited. Pleural tissue and fluid were collected by medical thoracoscopy and subjected to culture, Xpert MTB/RIF (Xpert) and Xpert-Ultra assays. Histopathological examination was also performed with the tissue and used as the major reference. Results: Sixty-one patients were enrolled, including: 27 tuberculosis (TB) pleurisy, 15 malignancy and 19 other chronic infection cases. The sensitivity, specificity, positive predictive value, and negative predictive value of Xpert-Ultra for TB diagnosis were 85.19% (23/27), 97.06% (33/34), 95.83% (23/24), and 89.19% (33/37), respectively. Xpert-Ultra testing with the biopsy tissue alone had an equivalent diagnostic capacity to that of pathological examination for the diagnosis of confirmed TB cases. By combining the pathological examination with Xpert-Ultra for biopsy, the percentage of confirmed TB cases greatly increased (i.e. 92.59% (25/27)). The “trace” positive outcome of Xpert-Ultra was highly supportive of TB diagnosis for both biopsy tissue and pleural fluid examinations. Conclusion: With the specimens collected by medical thoracoscopy, the Xpert-Ultra assay presented high value in identifying TB among pleurisy patients who had difficulties in etiological diagnosis.

Background: Linezolid presents strong antimicrobial activity against multidrug-resistant (MDR) pulmonary tuberculosis (TB), but its application in osteoarticular tuberculosis treatment remains understudied. Our objective was to analyze the bone penetration efficiency of linezolid in osteoarticular TB patients. Methods: Osteoarticular TB patients, treated with 600 mg q 24 h linezolid-containing regimens and undergoing surgery, were prospectively and consecutively enrolled. One dose linezolid was administered before surgery. Blood and bone samples were collected simultaneously during operation, and their linezolid concentrations were then detected using high-performance liquid chromatography-tandem mass spectrometry. Pus samples were subjected to mycobacterial culture and GeneXpert MTB/RIF assay. The minimum inhibition concentrations (MICs) and drug susceptibility testing were performed with the recovered isolates. Results: A total of 36 eligible osteoarticular TB patients were enrolled, including five MDR/rifampicin-resistant cases. All the 12 recovered isolates had MICs ≤0.5 μg/mL for linezolid. Mean concentrations in plasma, collected 100−510 min after the preoperative dosing, were 10.43 ± 4.83 μg/mL (range 3.29−22.26 μg/mL), and median concentrations in bone were 3.93 μg/mL (range 0.61−16.34 μg/mL). The median bone/plasma penetration ratio was 0.42 (range 0.14−0.95 μg/mL). Linezolid concentration in bone had a linear correlation with the drug concentration in plasma (r = 0.7873, p < 0.0001), while plasma concentration could explain 61.98% of the variation of concentration in bone (R2 = 0.6198). Notably, stratification analysis by sampling time demonstrated that samples collected 200−510 min after dosing had very good linear relationships between their bone and plasma concentrations (r = 0.9323). Conclusions: Linezolid penetrates from blood to bone efficiently, and the penetration further stabilizes ∼3 h after dosing.

Antibody responses against Mycobacteria infection have been reported, but whether and how they impact anti-bacteria immunity in the host is unclear. Here the authors characterize human anti-Mycobacteria antibodies to find them targeting a Mycobacteria transporter protein, PstS1, show distinct interaction modes in crystal structure, and mediate protection in vitro.

More sensitive, rapid, and affordable diagnostic tools for pulmonary tuberculosis (PTB) are urgently needed. This study aimed to assess the performance of EasyNAT MTC (abbreviation: EasyNAT) (Ustar Biotechnologies, China), a novel isothermal amplification method with a turnaround time of less than two hours that requires a few manual steps to process the sputum. Sputum samples from 249 patients with suspected PTB were subjected to smear, culture, Xpert MTB/RIF (Cepheid, USA) and EasyNAT assay testing. Of the 169 PTB patients, EasyNAT detected more PTB patients than Xpert (72.19% vs. 61.54%, P

Abstract Introduction To evaluate the value of the combination of computed tomography (CT)-guided percutaneous lung biopsy and Xpert MTB/RIF Ultra (Xpert-Ultra) in enhancing the differential diagnosis of tuberculosis. Methods Patients with lesions in the lungs for whom the differential diagnosis was difficult were prospectively and consecutively enrolled. Specimens collected by percutaneous lung biopsy were subject to Xpert-Ultra and histopathologic examination, respectively. Results Biopsy was successfully performed for 147 patients who were eligible for analysis, including 65 TB, 55 lung cancer and 27 other chronic infection cases. The sensitivity, specificity, positive predictive value and negative predictive value of Xpert-Ultra for TB diagnosis were 75.38% (49/65) and 95.12% (78/82), 92.45% (49/53) and 82.98% (78/94), respectively. Among patients with strong evidence for TB diagnosis (categorized as confirmed or probable TB cases), the overall positive rate of the Xpert-Ultra assay was 83.63% (46/55), which was much higher than for any reported sample type with negative smear test outcomes to date. Five rifampicin (RIF)-resistant cases were identified using the biopsy tissues. However, “trace” positive did not seem reliable for TB diagnosis with lung biopsy specimens; a 25.61% (21/82) “trace”-positive rate was acquired in the non-TB group. Conclusion Percutaneous lung biopsy combined with Xpert-Ultra produced high sensitivity for culture-negative pulmonary TB patient diagnosis. “Trace” outcome might not be a reliable positive category for lung biopsy specimens. Trial registration ClinicalTrials.gov identifier, ChiCTR1900026412. Retrospectively registered on Oct 8th, 2019).

Abstract Background The integrity of cell wall structure is highly significant for the in vivo survival of mycobacteria. We hypothesized that changes in morphology may indicate changes in cell wall metabolism and identified an aceE gene mutant (aceE-mut) which presented a deficient colony morphology on 7H10 agar by screening transposon mutagenesis in Mycolicibacterium smegmatis, basonym Mycobacterium smegmatis (M. smegmatis). This study aimed to identify the functional role of aceE gene in cell wall biosynthesis in M. smegmatis. Results We observed that the colony morphology of aceE-mut was quite different, smaller and smoother on the solid culture medium than the wild-type (WT) strain during the transposon library screening of M. smegmatis. Notably, in contrast with the WT, which aggregates and forms biofilm, the aceE-mut lost its ability of growing aggregately and biofilm formation, which are two very important features of mycobacteria. The morphological changes in the aceE-mut strain were further confirmed by electron microscopy which indicated smoother and thinner cell envelope images in contrast with the rough morphology of WT strains. Additionally, the aceE-mut was more fragile to acidic stress and exhibited a pronounced defects in entering the macrophages as compared to the WT. The analysis of mycolic acid (MA) using LC-MS indicated deficiency of alpha-MA and epoxy-MA in aceE-mut strain whereas complementation of the aceE-mut with a wild-type aceE gene restored the composition of MA. Conclusions Over all, this study indicates that aceE gene plays a significant role in the mycolic acid synthesis and affects the colony morphology, biofilm formation of M. smegmatis and bacteria invasion of macrophage.

Sugarcane smut is the most severe sugarcane disease in China. The typical symptom is the emerging of a long, black whip from the top of the plant cane. However, in 2018, for the first time we observed the floral structures of sugarcane infected by smut fungus in the planting fields of China. Such smut-associated inflorescence in sugarcane was generally curved and short, with small black whips emerging from glumes of a single floret on the cane stalk. Compatible haploid strains, named Ssf1-7 (MAT-1) and Ssf1-8 (MAT-2), isolated from teliospores that formed black whips in inflorescence of sugarcane were selected for sexual mating assay, ITS DNA sequencing analysis and pathogenicity assessment. The isolates Ssf1-7 and Ssf1-8 showed stronger sexual mating capability than the reported Sporisorium scitamineum strains Ss17 and Ss18. The ITS DNA sequence of the isolates Ssf1-7 and Ssf1-8 reached 100% similarity to the isolates of S. scitamineum strains available in GenBank. Inoculating Ssf1-7 + Ssf1-8 to six sugarcane varieties, i.e., GT42, GT44, GT49, GT55, LC05-136 and ROC22, resulted in different smut morphological modifications. The symptoms of floral structure only occurred in LC05-136, indicating that the flowering induction by S. scitamineum is variety-specific. Furthermore, six selected flowering-related genes were found to be differentially expressed in infected Ssf1-7 + Ssf1-8 LC05-13 plantlets compared to uninfected ones. It is concluded that the flowering induction by S. scitamineum depends on specific fungal race and sugarcane variety, suggesting a specific pathogen–host interaction and expression of some flowering-related genes.

Background: Tuberculosis recurrence is still a major problem for the control of tuberculosis, and the cause of the recurrence is still unclear.Methods: We retrospectively recruited 68 pairs of samples of Mycobacterium tuberculosis (MTB) from recurrent TB cases in Beijing Chest Hospital between January 2008 and December 2019. The whole-genome sequencing was conducted to analyze single-nucleotide polymorphism (SNP) and to identify whether recurrent disease was due to relapse or reinfection. The BACTEC MGIT was performed to compare differences in drug susceptibility profiles between two episodes.Results: 62 (91.2%) out of 68 confirmed recurrence were due to relapse, whereas the remaining six (8.8%) were due to reinfection. And there was a strong association between earlier relapse and underlying chronic diseases. In addition, the MTB isolates from non-diabetic patients had a higher mutation rate than those from diabetic patients. A community transmission was also identified in our cohort. Levofloxacin resistance was the most frequently observed drug resistance for 12.9% relapse cases.Conclusion: The relapse of a previous episode in Beijing. The underlying chronic diseases are associated with an earlier TB relapse. MTB isolates were more prone to develop levofloxacin resistance than moxifloxacin resistance after FQ exposure. The patients at high-risk for relapses deserve more careful investigation.

Arsenic can cause neurodegenerative diseases of the brain, but the definite mechanism is still unknown. In this study, to discuss the disturbances on brain metabolome and lipidome under subchronic arsenic exposure, we treated mice with the arsenic-containing feed (concentration of total arsenic = 30 mg/kg) prepared in accordance with the proportion of rice arsenicals for 16 weeks and performed metabolomics and lipidomics studies respectively using UHPLC-Triple-TOF-MS/MS and UHPLC-Q Exactive Focus MS/MS on mice brain. In addition, the distributions of arsenical metabolites along the feed-gut-blood-brain chain were analyzed by ICP-MS and HPLC-ICP-MS, and fecal microbial variations were investigated by 16 s sequencing. The data showed that although only a tiny amount of arsenic (DMA=0.101 mg/kg, uAs=0.071 mg/kg) enters the brain through the blood-brain barrier, there were significant changes in brain metabolism, including 118 metabolites and 17 lipids. These different metabolites were involved in 30 distinct pathways, including glycometabolism, and metabolisms of lipid, nucleic acid, and amino acid were previously reported to be correlated with neurodegenerative diseases. Additionally, these different metabolites were significantly correlated with 12 gut bacterial OTUs, among which Lachnospiraceae, Muribaculaceae, Ruminococcaceae, and Erysipelotrichaceae were also previously reported to be related to the distortion of metabolism, indicating that the disturbance of metabolism in the brain may be associated with the disturbance of gut microbes induced by arsenic. Thus, the current study demonstrated that the brain metabolome and lipidome were significantly disturbed under subchronic arsenic exposure, and the disturbances also significantly correlated with some gut microbiome and may be associated with neurodegenerative diseases. Although preliminary, the results shed some light on the pathophysiology of arsenic-caused neurodegenerative diseases.

In China, sugarcane (Saccharum spp.) hybrid cross-breeding began in 1953; approximately 70 years since then, >100 commercial sugarcane varieties have been created. In this study, 88 commercial varieties bred in China between 1953 and 2010 and 12 original foundational varieties were planted to investigate the effect of improving sugarcane varieties in China. Considering 20 years as a time node, the commercial varieties were classified into four improved generations. Retrospective analysis showed significant improvements in sucrose and other technological characteristics of commercial sugarcane varieties. The adoption of improved varieties over generations has continuously increased sugarcane’s sucrose, juice sugar, and gravity purity, and the difference was significant between Gen1 and Gen3, and between Gen2 and Gen4. Gen4 showed 2.06%, 2.35%, and 3.69% higher sugarcane sucrose (p < 0.01), juice sugar (p < 0.01), and purity (p < 0.05), respectively, and 1.13% lower sugarcane fiber (p < 0.01) than Gen1, the original foundational hybrid varieties. The development of new varieties has improved the technological characteristics of Chinese sugarcane. Sugarcane sucrose, juice sugar, and purity showed an increasing trend. Sugarcane fiber content did not significantly change with the development of new varieties but declined in comparison with the original foundational hybrid varieties.

Abstract Background Isoniazid (INH) represents the cornerstone for the treatment of cases infected with Mycobacterium tuberculosis (MTB) strains. Several molecular mechanisms have been shown to be the major causes for INH resistance, while the dynamic change of mutations conferring INH resistance among MTB strains during the past decade is still unknown in China. Methods In this study, we carried out a comparative analysis of the INH minimal inhibitory concentration (MIC) distribution, and investigate the dynamic change of molecular characteristics among INH-resistant MTB strains between 2005 and 2015. Results The proportion of INH resistance (39.0%, 105/269) in 2015 was significantly higher than in 2005 (30.0%, 82/273; P = 0.03). Among 269 isolates collected in 2015, 76 (28.3%, 76/269) exhibited high-level INH-resistance (MIC≥32 mg/L), which was significantly higher than that in 2005 (20.5%, 56/273, P = 0.04). In addition, a significantly higher percentage of INH-resistant isolates carried inhA promoter mutations in 2015 (26.7%) versus that in 2005 (14.6%, P = 0.04), while no significant difference was observed in the rates of isolates containing katG mutations between 2005 (76.8%) and 2015 (70.5%, P = 0.33). Notably, the proportion of MTB isolates with inhA mutations (26.7%, 28/105) for patients who had previous exposure to protionamide (PTH) was higher than that for patients who had no previous exposure to PTH (21.4%, 6/28). Conclusions In conclusion, our results demonstrated that the proportion of INH-resistant MTB isolates significantly increased during the last decade, which was mainly attributed to an increase of high-level INH-resistant MTB. In addition, prior exposure to PTH may be associated with the increased frequency of INH-resistant tuberculosis strains with inhA mutations in China.

We investigated the epidemiology of extrapulmonary tuberculosis (TB) among patients admitted to Beijing Chest Hospital, Beijing, China, during January 2008–December 2017. Of 19,279 hospitalized TB patients, 33.4% (6,433) had extrapulmonary TB and 66.6% (12,846) had pulmonary TB. The most frequent forms of extrapulmonary TB observed were skeletal TB (41.1%) and pleural TB (26.0%). Younger, female patients from rural areas were more likely to have extrapulmonary TB. However, patients with diabetes mellitus were less likely to have extrapulmonary TB compared with patients without diabetes. A higher proportion of multidrug-resistant (MDR) TB was observed among patients with extrapulmonary TB than among patients with pulmonary TB. We observed a large increase in MDR TB, from 17.3% to 35.7%, for pleural TB cases. The increasing rate of drug resistance among extrapulmonary TB cases highlights the need for drug susceptibility testing and the formulation of more effective regimens for extrapulmonary TB treatment.

The aim of this study was to develop a simple and reliable method to detect Mycobacterium tuberculosis complex (MTBC) and verify its clinical application preliminarily. A loop-mediated isothermal amplification method coupled with lateral flow biosensor (LAMP-LFB) assay, was developed and evaluated for detection of MTBC. Two sets of primers, which targeted IS6110 and IS1081 sequences of MTBC, were designed for establishment of multiplex LAMP-LFB assay. The amplicons were labelled with biotin and fluorescein isothiocyanate (FITC) by adding FITC labelled primer and biotin-14-dATP and biotin-14-dCTP and could be visualized using LFB. The optimal reaction conditions of multiplex LAMP-LFB assay confirmed were 66°C for 50 min. The analytical sensitivity of multiplex LAMP-LFB is 10 fg of genomic templates using pure culture, and no cross-reactivity with other common bacteria and non-tuberculous mycobacteria strains was obtained. A total of 143 clinical samples collected from 100 TB patients (62 definite TB cases and 38 probable TB cases) and 43 non-TB patients were used for evaluating the feasibility of multiplex LAMP-LFB assay. The multiplex LAMP-LFB (82.0%, 82/100) showed higher sensitivity than culture (47.0%, 47/100, P < 0.001) and Xpert MTB/RIF (54.0%, 54/100, P < 0.001). Importantly, the multiplex LAMP-LFB assay detected additional 28 probable TB cases, which increased the percentage of definite TB cases from 62.0% (62/100) to 90.0% (90/100). The specificity of multiplex LAMP-LFB assay in patients without TB was 97.7% (42/43). Therefore, multiplex LAMP-LFB assay is a simple, reliable, and sensitive method for MTBC detection, especially in probable TB cases and resource limited settings.

Pet ownership is an essential environmental exposure that might influence the health of the owner. This study's primary objectives were to explore the effects of cat ownership on the gut microbial diversity and composition of owners. Raw data from the American Gut Project were obtained from the SRA database. A total of 214 Caucasian individuals (111 female) with cats and 214 individuals (111 female) without cats were used in the following analysis. OTU number showed significant alteration in the Cat group and Female_cat group, compared with that of the no cat (NC) group and Female_ NC group, respectively. Compared with the NC group, the microbial phylum Proteobacteria was significantly decreased in the Cat group. The microbial families Alcaligenaceae and Pasteurellaceae were significantly reduced, while Enterobacteriaceae and Pseudomonadaceae were significantly increased in the Cat group. Fifty metabolic pathways were predicted to be significantly changed in the Cat group. Twenty-one and 13 metabolic pathways were predicted to be significantly changed in the female_cat and male_cat groups, respectively. Moreover, the microbial phylum Cyanobacteria was significantly decreased, while the families Alcaligenaceae, Pseudomonadaceae and Enterobacteriaceae were significantly changed in the normal weight cat group. In addition, 41 and 7 metabolic pathways were predicted to be significantly changed in the normal-weight cat and overweight cat groups, respectively. Therefore, this study demonstrated that cat ownership could influence owners' gut microbiota composition and function, especially in the female group and normal-weight group.

Betel nut chewing (BNC) is prevalent in South Asia and Southeast Asia. BNC can affect host health by modulating the gut microbiota. The aim of this study is to evaluate the effect of BNC on the gut microbiota of the host. Feces samples were obtained from 34 BNC individuals from Ledong and Lingshui, Hainan, China. The microbiota was analyzed by 16S rRNA gene sequencing. BNC decreased the microbial α-diversity. Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria were the predominant phyla, accounting for 99.35% of the BNC group. The Firmicutes-to-Bacteroidetes ratio was significantly increased in the BNC group compared to a control group. The abundances of the families Aerococcaceae, Neisseriaceae, Moraxellaceae, Porphyromonadaceae, and Planococcaceae were decreased in the BNC/BNC_Male/BNC_Female groups compared to the control group, whereas the abundances of Coriobacteriaceae, Streptococcaceae, Micrococcaceae, Xanthomonadaceae, Coxiellaceae, Nocardioidaceae, Rhodobacteraceae, and Succinivibrionaceae were increased. In general, the gut microbiome profiles suggest that BNC may have positive effects, such as an increase in the abundance of beneficial microbes and a reduction in the abundance of disease-related microbes. However, BNC may also produce an increase in the abundance of disease-related microbes. Therefore, extraction of prebiotic components could increase the beneficial value of betel nut.