Transmission dynamics of tuberculosis in a high-burden area of China: An 8-year population-based study using whole genome sequencing.

OBJECTIVES: This study investigated the transmission patterns of tuberculosis (TB) and its associated risk factors in Hunan province to inform the development of prevention and control strategies in the region. METHODS: An 8-year retrospective population-based genomic epidemiological study was conducted. Genomic clusters were defined using distance thresholds of 12-single-nucletide-polymorphisms. Risk factors associated with TB transmission were analyzed using logistic regression model. Kernel Density analysis was used to locate hotspots where transmission occurred. RESULTS: Among 2649 TB cases included in this study, 275 clusters were identified, with an overall clustering rate of 24.7% (654/2649). Nearly 95% (620/654) of clustered strains were isolated from the same county. Of the 275 clusters, 23 (8.4%, 23/275) had differences in drug-resistant profiles, with FQs resistance mutations occurring most frequently (52.2%, 12/23). Multivariate analysis identified male TB patients, those aged 30-60 years, ethnic minorities, nonfarmers, retreated TB patients, and individuals infected with MDR/RR-TB as independent risk factors for TB transmission (P < 0.05). Kernel density analysis showed that among the 5 drug-resistant surveillance sites, Leiyang had the highest clustering rate, followed by Yongshun, Qidong, Hecheng, and Taojiang. CONCLUSION: Recent transmission in the region is predominantly occurring within counties. The risk factors related to TB transmission and the hotspots where transmission occurs can provide a scientific basis for the formulation of targeted TB prevention and control strategies.

Prevalence and antimicrobial susceptibility pattern of Mycobacterium abscessus complex isolates in Chongqing, Southwest China.

Objectives: To investigate the prevalence of Mycobacterium abscessus complex (MABC), drug resistance characteristics, and the relationship between clarithromycin (CLA) susceptibility and MABC genotype in Chongqing, China. Methods: A total of 434 NTM patient isolates were collected between October 2018 and October 2019. Isolates confirmed to be non-tuberculous mycobacteria (NTM) were tested for minimal inhibitory concentrations of antimicrobial agents. In addition, rrl and erm(41) gene sequences were used to analyze the acquired macrolide resistance and inducible macrolide resistance. Results: Overall, 17 different NTM species were detected, of which M. abscessus (22.6 %, 91/403) was most prevalent. Amikacin, CLA, azithromycin and cefoxitin exhibited potent activities against MABC organisms, but no significant differences were observed in drug resistance rates between M. abscessus and M. massiliense (P > 0.05). On day 3 of culture, the acquired resistance rate against CLA was 7.4 % (9/121). Of 41 MABC isolates with inducible CLA resistant, 95.1 % (39/41) isolates belonged to the erm(41) T28 sequevar, while the remaining 4.9 % (2/41) possessed the M. massiliense genotype. All erm(41) C28 sequevar isolates were sensitive to CLA on day3 and day 14 of culture. Meanwhile, of the 5 erm(41) T28 isolates with acquired resistance, all possessed rrl 2058/2059 mutations, including 3 isolates with A2058C mutation and 2 isolates with A2059G mutation. While 2 of the 4 M. massiliense isolates with acquired resistance possessed the A2059G mutation, and one isolate possessed the A2058G mutation. Conclusion: Erm(41) and rrl gene could serve as useful markers for predicting macrolide susceptibility of MABC complex isolates.

A rare family outbreak of Mycobacterium abscessus infection in immunocompetent fraternal triplets.

Background: Mycobacterium abscessus (M. abscessus) infection is rare in children who were previously healthy, particularly in infants. We present the first report of a family outbreak of M. abscessus infection among immunocompetent infant triplets. Methods: We reviewed triplets' demographic data, laboratory tests and imaging examinations to describe their clinical features. We performed whole-exome sequencing to rule out primary immunodeficiency disorders. We used DNA sequencing for M. abscessus subspecies identification. Results: The fraternal triplets (triples A, B and C) presented with a 10-day history of cough. Triple A also experienced a brief episode of fever, and triple B had tachypnea. Chest CT scans showed pulmonary masses and nodules in triples A and C, and cavities in triple B. Cultures of sputum and bronchoalveolar lavage fluid from all triplets yielded M. abscessus. Further subspecies identification showed that isolates from triples A and C were M. abscessus subsp. massiliense, and isolates from triple B were M. abscessus subsp. abscessus (MAA). After eight months of combination therapy, the pulmonary lesions of the triplets improved significantly. Conclusion: Our study confirms that M. abscessus pulmonary disease can occur in immunocompetent infants. We hypothesize that the simultaneous infection of the triplets may be associated with their prematurity and extensive environmental exposure. This study highlights the importance to include M. abscessus infection in the differential diagnosis of pulmonary masses and/or cavities, regardless of the age of onset or the presence of underlying pathology or susceptible genes.

Systemic immune-inflammation index and serum glucose-potassium ratio predict poor prognosis in patients with spontaneous cerebral hemorrhage: An observational study.

Recent studies have shown systemic inflammatory response, serum glucose, and serum potassium are associated with poor prognosis in spontaneous intracerebral hemorrhage (SICH). This retrospective study aimed to investigate the association of systemic immune-inflammatory index (SII) and serum glucose-potassium ratio (GPR) with the severity of disease and the poor prognosis of patients with SICH at 3 months after hospital discharge. We reviewed the clinical data of 105 patients with SICH, assessed the extent of their disease using Glasgow Coma Scale score, National Institutes of Health Stroke Scale (NIHSS) score, and hematoma volume, and categorized them into a good prognosis group (0-3 scores) and a poor prognosis group (4-6 scores) based on their mRS scores at 3 months after hospital discharge. Demographic characteristics, clinical, laboratory, and imaging data at admission were compared between the 2 groups, bivariate correlations were analyzed using Spearman's correlation coefficients, multivariate logistic regression analysis was used to determine the independent risk factors for poor prognosis of patients with SICH, and finally, SII, GPR, and platelet/lymphocyte ratio (PLR) were examined using the subject's work characteristics (ROC) curve, lymphocyte/monocyte ratio (LMR), and neutrophil/lymphocyte ratio (NLR) for their predictive efficacy for poor prognosis. Patients in the poor prognosis group had significantly higher SII and serum GPR than those in the good prognosis group, and Spearman analysis showed that SII and serum GPR were significantly correlated with the admission Glasgow Coma Scale score as well as the NIHSS score and that SII and GPR increased with the increase in mRS score. Multivariate logistic regression analysis showed that admission NIHSS score, hematoma volume SII, GPR, NLR, and PLR were independently associated with poor patient prognosis. Analysis of the subjects' work characteristic curves showed that the areas under the SII, GPR, NLR, PLR, LMR, and coSII-GPR curves were 0.838, 0.837, 0.825, 0.718, 0.616, and 0.883. SII and GRP were significantly associated with disease severity and short-term prognosis in SICH patients 3 months after discharge, and SII and GPR had better predictive value compared with NLR, PLR, and LMR. In addition, coSII-GPR, a joint indicator based on SII and GPR, can improve the predictive accuracy of poor prognosis 3 months after discharge in patients with SICH.

Weed detection and recognition in complex wheat fields based on an improved YOLOv7.

Introduction: The precise detection of weeds in the field is the premise of implementing weed management. However, the similar color, morphology, and occlusion between wheat and weeds pose a challenge to the detection of weeds. In this study, a CSCW-YOLOv7 based on an improved YOLOv7 architecture was proposed to identify five types of weeds in complex wheat fields. Methods: First, a dataset was constructed for five weeds that are commonly found, namely, Descurainia sophia, thistle, golden saxifrage, shepherd's purse herb, and Artemisia argyi. Second, a wheat weed detection model called CSCW-YOLOv7 was proposed to achieve the accurate identification and classification of wheat weeds. In the CSCW-YOLOv7, the CARAFE operator was introduced as an up-sampling algorithm to improve the recognition of small targets. Then, the Squeeze-and-Excitation (SE) network was added to the Extended Latent Attention Networks (ELAN) module in the backbone network and the concatenation layer in the feature fusion module to enhance important weed features and suppress irrelevant features. In addition, the contextual transformer (CoT) module, a transformer-based architectural design, was used to capture global information and enhance self-attention by mining contextual information between neighboring keys. Finally, the Wise Intersection over Union (WIoU) loss function introducing a dynamic nonmonotonic focusing mechanism was employed to better predict the bounding boxes of the occluded weed. Results and discussion: The ablation experiment results showed that the CSCW-YOLOv7 achieved the best performance among the other models. The accuracy, recall, and mean average precision (mAP) values of the CSCW-YOLOv7 were 97.7%, 98%, and 94.4%, respectively. Compared with the baseline YOLOv7, the improved CSCW-YOLOv7 obtained precision, recall, and mAP increases of 1.8%, 1%, and 2.1%, respectively. Meanwhile, the parameters were compressed by 10.7% with a 3.8-MB reduction, resulting in a 10% decrease in floating-point operations per second (FLOPs). The Gradient-weighted Class Activation Mapping (Grad-CAM) visualization method suggested that the CSCW-YOLOv7 can learn a more representative set of features that can help better locate the weeds of different scales in complex field environments. In addition, the performance of the CSCW-YOLOv7 was compared to the widely used deep learning models, and results indicated that the CSCW-YOLOv7 exhibits a better ability to distinguish the overlapped weeds and small-scale weeds. The overall results suggest that the CSCW-YOLOv7 is a promising tool for the detection of weeds and has great potential for field applications.

Evaluation of genetic correlation with fluoroquinolones resistance in rifampicin-resistant Mycobacterium tuberculosis isolates.

Objective: To detect levofloxacin (LFX) and moxifloxacin (MFX) resistance among rifampicin-resistant tuberculosis (RR-TB) isolates, and predict the resistance level based on specific mutations in gyrA and gyrB genes. Methods: A total of 686 RR-TB isolates were collected from Chinese Drug Resistance Surveillance Program from 2013 to 2020. The minimum inhibitory concentrations (MICs) of 12 anti-TB drugs were acquired using the broth microdilution method, followed by whole genome sequencing (WGS) analysis. Results: Among the 686 RR isolates, the most prevalent resistance was to isoniazid (80.5 %) and ethambutol (28.4 %), followed by LFX (26.1 %) and MFX (21.9 %). The resistance rate of LFX (26.1%-99.4 %) was higher than that of MFX (21.9%-83.3 %) across various drug resistance patterns. Of the 180 fluoroquinolones (FQs) resistant isolates, 168 (93.3 %) had mutations in quinolone-resistant determining regions (QRDRs) with 21 mutation types, and Asp94Gly (32.7 %, 55/168) was the predominant mutation. Isolates with mutations in Asp94Asn and Asp94Gly were associated with high levels of resistance to LFX and MFX. Using broth microdilution method as gold standard, the sensitivities of WGS for LFX and MFX were 93.3 % and 98.0 %, and the specificities were 98.6 % and 95.0 %, respectively. Conclusion: The resistance rate of LFX was higher than that of MFX among various drug resistance patterns in RR-TB isolates. The gyrA Asp94Gly was the predominant mutation type underlying FQs resistance. However, no significant difference was observed between mutation patterns in gyrA gene and resistance level of FQs.

A pediatric case of dupilumab-induced pustular psoriasis.

Dupilumab is a novel treatment agent for moderate to severe atopic dermatitis (AD) with few adverse effects. Drug-induced psoriasiform lesions are rare.We report a 4-year-old boy with AD who developed pustular psoriasis during treatment with dupilumab.Pustular psoriasis appeared within 1 week of treatment and worsened in the second week. After stopping dupilumab administration, topical corticosteroids (desonide and mometasone furoate creams) and oral desloratadine without relief. Pustular psoriasis was confirmed by pathological examination, and thiamphenicol was administered. After 2 weeks of treatment, the lesions nearly resolved without recurrence in 1-year follow-up.Dupilumab-induced pustular psoriasis is rare in children.

Prevalence and genetic basis of Mycobacterium tuberculosis resistance to pretomanid in China.

BACKGROUND: Pretomanid is a key component of new regimens for the treatment of drug-resistant tuberculosis (TB) which are being rolled out globally. However, there is limited information on the prevalence of pre-existing resistance to the drug. METHODS: To investigate pretomanid resistance rates in China and its underlying genetic basis, as well as to generate additional minimum inhibitory concentration (MIC) data for epidemiological cutoff (ECOFF)/breakpoint setting, we performed MIC determinations in the Mycobacterial Growth Indicator Tube™ (MGIT) system, followed by WGS analysis, on 475 Mycobacterium tuberculosis (MTB) isolated from Chinese TB patients between 2013 and 2020. RESULTS: We observed a pretomanid MIC distribution with a 99% ECOFF equal to 0.5 mg/L. Of the 15 isolates with MIC values > 0.5 mg/L, one (MIC = 1 mg/L) was identified as MTB lineage 1 (L1), a genotype previously reported to be intrinsically less susceptible to pretomanid, two were borderline resistant (MIC = 2-4 mg/L) and the remaining 12 isolates were highly resistant (MIC ≥ 16 mg/L) to the drug. Five resistant isolates did not harbor mutations in the known pretomanid resistant genes. CONCLUSIONS: Our results further support a breakpoint of 0.5 mg/L for a non-L1 MTB population, which is characteristic of China. Further, our data point to an unexpected high (14/475, 3%) pre-existing pretomanid resistance rate in the country, as well as to the existence of yet-to-be-discovered pretomanid resistance genes.

Lapagyl mitigates UV-induced inflammation and immunosuppression via Foxp3+ Tregs and CCL pathway: A single-cell transcriptomics study.

BACKGROUND: As the largest organ of the body, the skin is constantly subjected to ultraviolet radiation (UVR), leading to inflammations and changes that mirror those seen in chronological aging. Although various small molecule drugs have been explored for treating skin photoaging, they typically suffer from low stability and a high incidence of adverse reactions. Consequently, the continued investigation of photoaging treatments, particularly those utilizing herbal products, remains a critical clinical endeavor. One such herbal product, Lapagyl, is derived from the bark of the lapacho tree and possesses antioxidant efficacies that could be beneficial in combating skin photoaging. PURPOSE: This research aimed to evaluate the efficacy of the herbal product Lapagyl in combating UVR-induced skin photoaging. Additionally, it sought to unravel the mechanisms by which Lapagyl promotes the regeneration of the skin extracellular matrix. METHODS: To investigate whether Lapagyl can alleviate skin aging and damage, a UVR radiation model was established using SKH-1 hairless mice. The dorsal skins of these mice were evaluated for wrinkle formation, texture, moisture, transepidermal water loss (TEWL), and elasticity. Pathological assessments were conducted to determine Lapagyl's efficacy. Additionally, single-cell sequencing and spectrum analysis were employed to elucidate the working mechanisms and primary components of Lapagyl in addressing UVR-induced skin aging and injury. RESULTS: Lapagyl markedly reduced UVR-induced wrinkles, moisture loss, and elasticity decrease in SKH-1 mice. Single-cell sequencing demonstrated that Lapagyl corrected the imbalance in cell proportions caused by UVR, decreased UVR-induced ROS expression, and protected basal and spinous cells from skin damage. Additionally, Lapagyl effectively prevented the entry of inflammatory cells into the skin by reducing CCL8 expression and curtailed the UVR-induced formation of Foxp3+ regulatory T cells (Tregs) in the skin. Both pathological assessments and ex vivo skin model results demonstrated that Lapagyl effectively reduced UVR-induced damage to collagen and elastin. Spectrum analysis identified Salidroside as the primary compound remaining in the skin following Lapagyl treatment. Taken together, our study elucidated the skin protection mechanism of the herbal product Lapagyl against UVR damage at the cellular level, revealing its immunomodulatory effects, with salidroside identified as the primary active compound for skin. CONCLUSION: Our study provided a thorough evaluation of Lapagyl's protective effects on skin against UVR damage, delving into the mechanisms at the cellular level. We discovered that Lapagyl mitigates skin inflammation and immunosuppression by regulating Foxp3+ Tregs and the CCL pathway. These insights indicate that Lapagyl has potential as a novel therapeutic option for addressing skin photoaging.

Evaluation of MeltPro Assay in Identification of Second-Line Injectable Drug Resistance in Multidrug-Resistant Tuberculosis Isolates.

Objective: We compared the MeltPro assay to whole-genome sequencing (WGS) to investigate the molecular characterization of second-line injectable drug (SLID) resistance in multidrug-resistant tuberculosis (MDR-TB) isolates in Chongqing, China. Methods: A total of 122 MDR-TB patient isolates were collected between March 2019 and June 2020 from Chongqing Municipality, China. Conventional drug-susceptibility testing was performed using the proportion method, followed to generate minimum inhibitory concentrations (MICs) of SLIDs determined by microplate alamarblue assay. All strains were subjected to both MeltPro and WGS assays. Results: Among 122 MDR-TB isolates, 30 (24.6%), 22 (18.0%), and 14 (11.5%) were resistant to kanamycin (KM), amikacin (AM), and capreomycin (CM), respectively. Of the 31 SLID-resistant isolates, 24 (77.4%, 24/31) isolates harbored mutations in the rrs gene, with the most prevalent mutations in rrs A1401G (22/24, 91.7%). Mutation in rrs A1401G was associated with high levels of resistance to KM (MIC, ≥40 µg/mL) and AM (MIC, ≥64 µg/mL), but disparities in CM-resistance levels. Using phenotypic drug-susceptibility testing as gold standard, we found that the overall sensitivity of MeltPro and WGS was 87.1% and 90.32% and specificity 100% and 97.8%, respectively. Seven isolates had discordant results between phenotypic and genotypic resistance of SLIDs. Conclusion: MeltPro is a promising diagnostic tool for accurate identification of SLID-resistant MTB isolates with mutations in the rrs and eis genes. There was a disparity between MeltPro with WGS results in the proportion of heterogeneous drug-resistant bacteria with rrs mutation and limited probes. Resistance mechanisms other than genetic mutations will affect the consistency of MeltPro and WGS with phenotypic drug-susceptibility results.

Transmission characteristics in Tuberculosis by WGS: nationwide cross-sectional surveillance in China.

China, with the third largest share of global tuberculosis cases, faces a substantial challenge in its healthcare system as a result of the high burden of multidrug-resistant and rifampicin-resistant tuberculosis (MDR/RR-TB). This study employs a genomic epidemiological approach to assess recent tuberculosis transmissions between individuals, identifying potential risk factors and discerning the role of transmitted resistant isolates in the emergence of drug-resistant tuberculosis in China. We conducted a population-based retrospective study on 5052 Mycobacterium tuberculosis (MTB) isolates from 70 surveillance sites using whole genome sequencing (WGS). Minimum spanning tree analysis identified resistance mutations, while epidemiological data analysis pinpointed transmission risk factors. Of the 5052 isolates, 23% (1160) formed 452 genomic clusters, with 85.6% (387) of the transmissions occurring within the same counties. Individuals with younger age, larger family size, new cases, smear positive, and MDR/RR were at higher odds for recent transmission, while higher education (university and above) and occupation as a non-physical workers emerged as protective factors. At least 61.4% (251/409) of MDR/RR-TB were likely a result of recent transmission of MDR/RR isolates, with previous treatment (crude OR = 2.77), smear-positive (cOR = 2.07) and larger family population (cOR = 1.13) established as risk factors. Our findings highlight that local transmission remains the predominant form of TB transmission in China. Correspondingly, drug-resistant tuberculosis is primarily driven by the transmission of resistant tuberculosis isolates. Targeted interventions for high-risk populations to interrupt transmission within the country will likely provide an opportunity to reduce the prevalence of both tuberculosis and drug-resistant tuberculosis.

Epidemiology of tuberculosis among children in Beijing, China, 2012-2021.

Data on epidemiology trends of paediatric tuberculosis (TB) are limited in China. So, we investigated the clinical and epidemiological profiles in diagnosed TB disease and TB infection patients at Beijing Children's Hospital. Of 3 193 patients, 51.05% had pulmonary TB (PTB) and 15.16% had extrapulmonary TB (EPTB). The most frequent forms of EPTB were TB meningitis (39.05%), pleural TB (29.75%), and disseminated TB (10.33%). PTB patients were significantly younger and associated with higher hospitalization frequency. Children aged 1-4 years exhibited higher risk of PTB and TB meningitis, and children aged 5-12 years had higher risk of EPTB. The proportion of PTB patients increased slightly from 40.9% in 2012 to 65% in 2019, and then decreased to 17.8% in 2021. The percentage of EPTB cases decreased from 18.3% in 2012 to 15.2% in 2019, but increased to 16.4% in 2021. Among EPTB cases, the largest increase was seen in TB meningitis. In conclusion, female and young children had higher risk of PTB in children. TB meningitis was the most frequent forms of EPTB among children, and young children were at high risk of TB meningitis. The distribution of different types of EPTB differed by age.

Apoptosis and pyroptosis in the nasal mucosa of Syrian hamster during SARS-CoV-2 infection and reinfection.

In SARS-CoV-2 infection, it has been observed that viral replication lasts longer in the nasal mucosa than in the lungs, despite the presence of a high viral load at both sites. In hamsters, we found that the nasal mucosa exhibited a mild inflammatory response and minimal pathological injuries, whereas the lungs displayed a significant inflammatory response and severe injuries. The underlying cellular events may be induced by viral infection in three types of cell death: apoptosis, pyroptosis, and necroptosis. Our findings indicate that apoptosis was consistently activated during infection in the nasal mucosa, and the levels of apoptosis were consistent with the viral load. On the other hand, pyroptosis and a few instances of necroptosis were observed only on 7 dpi in the nasal mucosa. In the lungs, however, both pyroptosis and apoptosis were prominently activated on 3 dpi, with lower levels of apoptosis compared to the nasal mucosa. Interestingly, in reinfection, obvious viral load and apoptosis in the nasal mucosa were detected on 3 dpi, while no other forms of cell death were detected. We noted that the inflammatory reactions and pathological injuries in the nasal mucosa were milder, indicating that apoptosis may play a role in promoting lower inflammatory reactions and milder pathological injuries and contribute to the generation of long-term viral replication in the nasal mucosa. Our study provides valuable insights into the differences in cellular mechanisms during SARS-CoV-2 infection and highlights the potential significance of apoptosis regulation in the respiratory mucosa for controlling viral replication.

Protocol for optimized dissociation of human scalp tissue for hair follicle transcriptomics by scRNA-seq.

Single-cell RNA sequencing (scRNA-seq) is a powerful tool for studying transcriptomics. Here, we present an optimized protocol for dissociating human scalp tissue and acquiring high-quality single-cell suspension for scRNA-seq to study transcriptomics of human hair follicles. We describe steps for human scalp tissue cleaning, subcutaneous fat removal, mechanical mincing, and enzymatic digestion. We then detail procedures for cleaning, resuspending, a cell viability assay, and library construction.

Association of Interleukin-17A and Interleukin-17F Gene Polymorphisms with Atopic Dermatitis in Chinese Children.

Background: Atopic dermatitis (AD) is a chronic, recurrent inflammatory disease associated with an unbalanced immune response in the upper layers of the skin tissue, mostly starting in childhood. As important factors in gene expression regulation, polymorphisms in interleukin (IL)-17A and IL-17F may be associated with the susceptibility and severity of AD. Methods: Blood samples and clinical information were obtained from 132 patients with AD and 100 healthy children. Using multiplex polymerase chain reaction and next-generation sequencing, five potential single-nucleotide polymorphisms (SNPs) of IL-17A and IL-17F were genotyped in all participants. The relationship between SNPs and susceptibility to or severity of AD was examined by analyzing haplotypes and genetic models. Results: The IL-17A rs3819025 polymorphism was substantially associated with higher AD risk in both the allele model (p = 0.03; odds ratio [OR] = 1.76; confidence interval [CI]: 1.05-2.95) and the dominant model (p = 0.04, OR = 1.85; CI: 1.03-3.33). There was no correlation between AD susceptibility and the IL-17A (rs2275913 and rs4711998) or IL-17F (rs763780 and rs12203736) SNPs (all p > 0.05). Additionally, the five IL-17A and IL-17F SNPs did not significantly differ across the mild-to-moderate and severe subgroups (all p > 0.05). Conclusions: The IL-17A/rs3819025 polymorphism was linked to the development of AD, whereas the IL-17F polymorphism was unrelated to the susceptibility to and severity of AD. The IL-17A polymorphism may provide valuable information to speculate on the susceptibility to AD in Chinese Han children.

The Abundant Distribution and Duplication of SARS-CoV-2 in the Cerebrum and Lungs Promote a High Mortality Rate in Transgenic hACE2-C57 Mice.

Patients with COVID-19 have been reported to experience neurological complications, although the main cause of death in these patients was determined to be lung damage. Notably, SARS-CoV-2-induced pathological injuries in brains with a viral presence were also found in all fatal animal cases. Thus, an appropriate animal model that mimics severe infections in the lungs and brain needs to be developed. In this paper, we compared SARS-CoV-2 infection dynamics and pathological injuries between C57BL/6Smoc-Ace2em3(hACE2-flag-Wpre-pA)Smoc transgenic hACE2-C57 mice and Syrian hamsters. Importantly, the greatest viral distribution in mice occurred in the cerebral cortex neuron area, where pathological injuries and cell death were observed. In contrast, in hamsters, viral replication and distribution occurred mainly in the lungs but not in the cerebrum, although obvious ACE2 expression was validated in the cerebrum. Consistent with the spread of the virus, significant increases in IL-1ß and IFN-γ were observed in the lungs of both animals. However, in hACE2-C57 mice, the cerebrum showed noticeable increases in IL-1ß but only mild increases in IFN-γ. Notably, our findings revealed that both the cerebrum and the lungs were prominent infection sites in hACE2 mice infected with SARS-CoV-2 with obvious pathological damage. Furthermore, hamsters exhibited severe interstitial pneumonia from 3 dpi to 5 dpi, followed by gradual recovery. Conversely, all the hACE2-C57 mice experienced severe pathological injuries in the cerebrum and lungs, leading to mortality before 5 dpi. According to these results, transgenic hACE2-C57 mice may be valuable for studying SARS-CoV-2 pathogenesis and clearance in the cerebrum. Additionally, a hamster model could serve as a crucial resource for exploring the mechanisms of recovery from infection at different dosage levels.

Diagnostic efficacy of an optimized nucleotide MALDI-TOF-MS assay for anti-tuberculosis drug resistance detection.

PURPOSE: We aimed at evaluating the diagnostic efficacy of a nucleotide matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) assay to detect drug resistance of Mycobacterium tuberculosis. METHODS: Overall, 263 M. tuberculosis clinical isolates were selected to evaluate the performance of nucleic MALDI-TOF-MS for rifampin (RIF), isoniazid (INH), ethambutol (EMB), moxifloxacin (MXF), streptomycin (SM), and pyrazinamide (PZA) resistance detection. The results for RIF, INH, EMB, and MXF were compared with phenotypic microbroth dilution drug susceptibility testing (DST) and whole-genome sequencing (WGS), and the results for SM and PZA were compared with those obtained by WGS. RESULTS: Using DST as the gold standard, the sensitivity, specificity, and kappa values of the MALDI-TOF-MS assay for the detection of resistance were 98.2%, 98.7%, and 0.97 for RIF; 92.8%, 99%, and 0.90 for INH; 82.4%, 98.0%, and 0.82 for EMB; and 92.6%, 99.5%, and 0.94 for MXF, respectively. Compared with WGS as the reference standard, the sensitivity, specificity, and kappa values of the MALDI-TOF-MS assay for the detection of resistance were 97.4%, 100.0%, and 0.98 for RIF; 98.7%, 92.9%, and 0.92 for INH; 96.3%, 100.0%, and 0.98 for EMB; 98.1%, 100.0%, and 0.99 for MXF; 98.0%, 100.0%, and 0.98 for SM; and 50.0%, 100.0%, and 0.65 for PZA. CONCLUSION: The nucleotide MALDI-TOF-MS assay yielded highly consistent results compared to DST and WGS, suggesting that it is a promising tool for the rapid detection of sensitivity to RIF, INH, EMB, and MXF.

Deconstructing heterogeneity of replicative senescence in human mesenchymal stem cells at single cell resolution.

Following prolonged cell division, mesenchymal stem cells enter replicative senescence, a state of permanent cell cycle arrest that constrains the use of this cell type in regenerative medicine applications and that in vivo substantially contributes to organismal ageing. Multiple cellular processes such as telomere dysfunction, DNA damage and oncogene activation are implicated in promoting replicative senescence, but whether mesenchymal stem cells enter different pre-senescent and senescent states has remained unclear. To address this knowledge gap, we subjected serially passaged human ESC-derived mesenchymal stem cells (esMSCs) to single cell profiling and single cell RNA-sequencing during their progressive entry into replicative senescence. We found that esMSC transitioned through newly identified pre-senescent cell states before entering into three different senescent cell states. By deconstructing this heterogeneity and temporally ordering these pre-senescent and senescent esMSC subpopulations into developmental trajectories, we identified markers and predicted drivers of these cell states. Regulatory networks that capture connections between genes at each timepoint demonstrated a loss of connectivity, and specific genes altered their gene expression distributions as cells entered senescence. Collectively, this data reconciles previous observations that identified different senescence programs within an individual cell type and should enable the design of novel senotherapeutic regimes that can overcome in vitro MSC expansion constraints or that can perhaps slow organismal ageing.

Diagnostic value of tNGS vs Xpert MTB/RIF in childhood TB.

Objectives: To evaluate the diagnostic value of targeted next generation sequencing (tNGS) in childhood tuberculosis (TB) and compare the accuracy with Xpert MTB/RIF method. Methods: Children aged ≤18 years with symptoms suggestive of TB during July 2021 to December 2022 at Beijing Children's Hospital were included, and the performances of tNGS and Xpert were evaluated. Results: A total of 103 children with suspected TB were recruited, including 72 discharge diagnosis of TB and 31 non-TB cases. The mean age was 7.37 ± 4.77 years, and 62.1 % were male. The most common type of specimens was gastric aspirate (GA) (59, 57.3 %). Among all the 72 TB patients, tNGS showed higher sensitivity than Xpert, but the difference was not significant (34.7 %, 25/72 vs 20.8 %, 15/72; P = 0.063). The specificities of tNGS and Xpert were 87.1 % (27/31) and 96.8 % (30/31), respectively (P = 0.162). Among different types of specimen, the highest sensitivity of tNGS on sputum and pus was observed (80.0 %, 4/5), followed by pleural effusion (50.0 %, 2/4). One rifampin resistance and one protionamide resistance were detected in bacteriologically confirmed TB by tNGS. Conclusion: tNGS had a higher sensitivity but lower specificity compared to Xpert in diagnosis of children TB. tNGS yielded higher sensitivity than Xpert on gastric aspirate and sputum and pus.

Interferon-gamma release assay for screening of tuberculosis infection in children.

BACKGROUND: Interferon-gamma release assay (IGRA) is the main tool for the diagnosis of latent tuberculosis (TB) infection (LTBI). However, the indeterminate results were more frequent in children, and the underlying reasons were largely speculative. We aimed to compare QuantiFERON-TB Gold In-Tube (QFT-GIT) with X.DOT-TB (XDOT) for diagnosing LTBI, and to identify the risk factors associated with indeterminate results in children. METHODS: A retrospective study for children<18 years old, at risk for LTBI or progression to TB disease, received either QFT-GIT or X.DOT-TB tests was performed at Beijing Children's Hospital from August 2019 to August 2022. RESULTS: A total of 33,662 children were recruited, including 15,129 (44.9%) tested with X.DOT-TB and 18,533 (55.1%) with QFT-GIT. Proportion of positive and indeterminate results in children with respiratory disease was significantly higher than did that with other diseases, respectively (P < 0.001). The indeterminate rate of X.DOT-TB and QFT-GIT results decreased with increasing age (P < 0.001). Proportion of QFT-GIT indeterminate results was higher than that of X.DOT-TB across age groups. Male, age and disease classification all presented a statistically significant association with indeterminate IGRA results. CONCLUSIONS: The positive rates of X.DOT-TB and QFT-GIT in children were 3.1% and 1.8%, respectively. The X.DOT-TB assay performed better than QFT-GIT in children, and male, age and underlying diseases were associated with an increased risk of indeterminate IGRA results.