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Abstract Background Pantoea agglomerans (P. agglomerans) is a gram-negative bacterium that is commonly isolated from plant surfaces, seeds, and the environment. As an opportunistic pathogen, it can cause blood, urinary and soft tissue infections in immunocompromised patients. In central nervous system, P. agglomerans infection has been report in children and immune-compromised patients, however, infection by such bacterium in nontraumatized immune competent adults has not been reported. Here, we report a case of P. agglomerans cerebrospinal meningitis accompanied by positive anti-myeloperoxidase (MPO) antibody in a 49-year-old female who has a history of black fungus planting. Case presentation The patient manifested with repeated fever, headache, generalized muscle pain, and neurological defects. Cerebrospinal fluid (CSF) tests revealed a moderately elevated number of polymorphonuclear leukocytes (50–193 × 106/L), low glucose levels (0.54–2.44 mmo1/L), and extremely high protein content (2.42–25.42 g/L). Blood tests showed positive anti-myeloperoxidase antibodies lasting for 1.5 year before turning negative. Spine MRI showed thickening and enhancement of the whole spinal meninges. CSF metagenomic next-generation sequencing (mNGS) revealed 75,189 specific DNA reads of P. agglomerans. The patient underwent spinal laminectomy due to meningeal adhesions. Pathological results revealed fibrous tissue proliferation, inflammatory infiltration with focal necrosis and calcification in the dura mater. The patient was successfully treated with sufficient antibiotics at 1-year follow-up. Conclusions People should be alert to CNS infections caused by P. agglomerans which presented with relatively mild clinical symptoms at onset, especially for those who contucts relevant agricultural and forestry work. The CSF characterization of P. agglomerans meningitis is elevated multiple nuclei white blood cells, significantly reduced glucose content, and markedly increased protein level which may be related to the secondary spinal membrane adhesions.

Abstract This study presents an approach that integrates compressed sensing technology with two-dimensional hyperchaotic coupled Fourier oscillator systems (2D-HCFOS) to address the challenge of slow encryption speeds in agricultural unmanned aerial vehicles (UAVs). The primary challenge in enhancing encryption speed lies in the limited capacity inherent in traditional chaotic-based systems and the computational complexity of their processes. The 2D-HCFOS utilizes a complex two-dimensional hybrid chaotic system, which significantly enhances the security of agricultural UAV image data. Notably, the image encryption process is performed on a personal computer connected to the drone, ensuring efficient processing. By integrating advanced Fourier series and nonlinear coupled oscillators, the model surpasses existing chaotic-based methods, improving both the pseudo-randomness and robustness of encryption. Additionally, incorporating Bonouille functions into the discrete cosine transform (DCT) domain results in a sparser measurement matrix, which is essential for efficient encryption on personal computers. The effectiveness of 2D-HCFOS in securely encrypting agricultural drone images has been rigorously validated through simulations and analytical evaluations using sophisticated row, rotation, and matrix encryption techniques. The improved security performance is further verified by comparative analysis. Compared with other models, the Lyapunov index of 2D-HCFOS is 15.1039, and the sample entropy is 2.4987, indicating that it possesses superior chaotic performance and encryption reliability.

A one-step QuEChERS automated detection technology for 22 pesticides in cowpeas and mangoes was established based on ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The sample was extracted by acetonitrile containing 1% acetic acid, added with EN extraction salt (anhydrous Mg2SO4, sodium chloride, trisodium citrate dihydrate, disodium hydrogen citrate sesquihydrate) to remove water, and then purified with C18 and graphitized carbon black (GCB). The target analytes were separated by ACQUITY UPLC BEH C18 chromatography column, using 0.01% formic acid aqueous solution (containing 2 mmol/L ammonium formate)-0.01% formic acid methanol as the mobile phase, and determined in multiple reaction monitoring (MRM). The results showed that the 22 pesticides had good linearity in the range of 0.01~25.00 µg/L, with coefficient of determination (R2) higher than 0.994. The limits of detection (LOD, S/N=3) were 0.05~2 µg/kg, and the limits of quantification (LOQ, S/N=10) were 0.1~5 µg/kg. At the spiked levels of 1×LOQ, 2×LOQ, and 10×LOQ, the average recoveries for cowpeas were 70.9%~116.6%, 71.9%~118.3%, and 73.7%~113.7%, respectively, and for mangoes were 70.2%~114.6%, 71.2%~113.7%, and 74.3%~118.5%, respectively. The relative standard deviations for them were all within 20%. The method was successfully applied to the detection of 42 actual samples, and 33 samples detected pesticide residues. This method has high sensitivity and a satisfactory level of automation, effectively reducing human operation steps. It is suitable for the simultaneous detection and accurate quantification of 22 pesticides and their metabolites.

A method of ultra-high performance liquid chromatography quadrupole electrostatic field Orbitrap mass spectrometry (UHPLC-Q/Orbitrap MS) was developed for rapid screening illegal additives in diet food. The efficacy of three extraction solvents for methanol, acetonitrile and acetonitrile containing 1% formic acid were evaluated for the extraction of illegal additives from green plums and jellies. Ultimately, acetonitrile is selected to extract illegally additives under ultrasonication. In order to achieve good separation of the additives by UHPLC in diet foods, four mobile phases were compared, including methanol (0.1% formic acid)-water (0.1% formic acid and 5 mmol/L ammonium acetate), acetonitrile (0.1% formic acid)-water (0.1% formic acid and 5 mmol/L ammonium acetate), acetonitrile-water (5 mmol/L ammonium acetate), acetonitrile-water. The effects of mobile phases and acidity levels on the chromatographic behavior of the additives were investigated, and acetonitrile-water containing 5 mmol/L ammonium acetate shows the best performance under appropriate gradient. Then, the MS data were acquired in Full MS/dd-MS2 mode on UHPLC-Q/Orbitrap MS with the matrix-matched external standard method for quantification. Based on the self-constructed database of illegal additives in diet foods, one unknown additive is detected in a diet food containing green plums and jelly. By characterizing the molecular ion peak, secondary fragment ion information, and isotopic information, the unknown compound is identified to be 4-chlorobisacetophenetidin, also an illegal additive, which was verified by computation of density functional theory and analysis of the standard substance. To validate the accuracy and reliability of the method, the limits of detection (LODs), the limits of quantification (LOQs), linear ranges, recoveries, relative standard deviations (RSDs) and matrix effect (ME) for eight compounds were determined. The LODs and LOQs of deacetylbisacodyl and bisacodyl are 0.001 and 0.005 mg/kg, respectively. The LODs and LOQs of other compounds are 0.05 and 0.1 mg/kg, respectively. The correlation coefficient (R2) is higher than 0.999. All of the compounds exhibit varying degrees of MEs from moderate to weak, and the recoveries is in the range of 71.02%-103.01% with the RSDs of 0.87%-13.24%. The approach was applied to diet food containing green plums and jelly. The 4-chlorobisacodyl are detected in 30% of the green plums, suggesting that this method is suitable for the rapid qualitative and quantitative analysis of illegal additives in diet food. It is also demonstrated that the established screening process plays a positive role in identifying potential risk substances. The combination of quantum chemistry computation and mass spectrometry provides a novel strategy for the identification of unknown additives in food.

Abstract Background R2R3-MYB transcription factors belong to one of the largest gene subfamilies in plants, and they are involved in diverse biological processes. However, the role of R2R3-MYB transcription factor subfamily genes in the response of rice (Oryza sativa L.) to salt stress has been rarely reported. Results In this study, we performed a genome-wide characterization and expression identification of rice R2R3-MYB transcription factor subfamily genes. We identified a total of 117 R2R3-MYB genes in rice and characterized their gene structure, chromosomal location, and cis-regulatory elements. According to the phylogenetic relationships and amino acid sequence homologies, the R2R3-MYB genes were divided into four groups. qRT-PCR of the R2R3-MYB genes showed that the expression levels of 10 genes significantly increased after 3 days of 0.8% NaCl treatment. We selected a high expression gene OsMYB2-115 for further analysis. OsMYB2-115 was highly expressed in the roots, stem, leaf, and leaf sheath. OsMYB2-115 was found to be localized in the nucleus, and the yeast hybrid assay showed that OsMYB2-115 has transcriptional activation activity. Conclusion This result provides important information for the functional analyses of rice R2R3-MYB transcription factor subfamily genes related to the salt stress response and reveals that OsMYB2-115 may be an important gene associated with salt tolerance in rice.

Abstract Objectives To develop and validate a dual-energy CT (DECT)-based model for noninvasively differentiating between benign and malignant breast lesions detected on DECT. Materials and methods This study prospectively enrolled patients with suspected breast cancer who underwent dual-phase contrast-enhanced DECT from July 2022 to July 2023. Breast lesions were randomly divided into the training and test cohorts at a ratio of 7:3. Clinical characteristics, DECT-based morphological features, and DECT quantitative parameters were collected. Univariate analyses and multivariate logistic regression were performed to determine independent predictors of benign and malignant breast lesions. An individualized model was constructed. Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic ability of the model, whose calibration and clinical usefulness were assessed by calibration curve and decision curve analysis. Results This study included 200 patients (mean age, 49.9 ± 11.9 years; age range, 22–83 years) with 222 breast lesions. Age, lesion shape, and the effective atomic number (Zeff) in the venous phase were significant independent predictors of breast lesions (all p 0.05 by the Hosmer-Lemeshow test) and provided enhanced net benefits than simple default strategies within a wide range of threshold probabilities in both cohorts. Conclusion The DECT-based model showed a favorable diagnostic performance for noninvasive differentiation between benign and malignant breast lesions detected on DECT. Critical relevance statement The combination of clinical and morphological characteristics and DECT-derived parameter have the potential to identify benign and malignant breast lesions and it may be useful for incidental breast lesions on DECT to decide if further work-up is needed. Key Points It is important to characterize incidental breast lesions on DECT for patient management. DECT-based model can differentiate benign and malignant breast lesions with good performance. DECT-based model is a potential tool for distinguishing breast lesions detected on DECT. Graphical Abstract

Abstract Aedes aegypti is a primary vector for transmitting various arboviruses, including Yellow fever, dengue and Zika virus. The mosquito midgut is the principal organ for blood meal digestion, nutrient absorption and the initial site of arbovirus infection. Although a previous study delineated midgut’s transcriptome of Ae. aegypti at the single-nucleus resolution, there still lacks an established protocol for isolating and RNA sequencing of single cells of Ae. aegypti midgut, which is required for investigating arbovirus-midgut interaction at the single-cell level. Here, we established an atlas of the midgut cells for Ae. aegypti by single-cell RNA sequencing. We annotated the cell clusters including intestinal stem cells/enteroblasts (ISC/EB), cardia cells (Cardia), enterocytes (EC, EC-like), enteroendocrine cells (EE), visceral muscle (VM), fat body cells (FBC) and hemocyte cells (HC). This study will provide a foundation for further studies of arbovirus infection in mosquito midgut at the single-cell level.

We conducted a large-scale survey of arboviruses in mosquitoes and biting midges to assess the presence and spread of mosquito-borne pathogens currently circulating on Hainan Island, China. RT-PCR assays were used to detect the arbovirus species, distribution, and infection rates in mosquitoes and biting midges. Cell inoculation and high throughput sequencing were performed to isolate the viruses and assemble full viral genomes. Phylogenetic analysis was conducted to identify the viral genotypes and evolutionary relationships with known viruses. During 2019-2023, 32,632 mosquitoes and 21,000 biting midges were collected from 14 of 18 cities/counties on Hainan Island. Japanese encephalitis virus (JEV) was detected in Culex mosquitoes from five cities/counties, where the minimum infection rate (MIR) was 1.6 (0.6-2.6) per 1,000 females tested. Tembusu virus (TMUV) was detected in Culex mosquitoes from three cities/counties with MIR1.0 (0-2.2) per 1,000. Getah virus (GETV) was detected in Armigeres mosquitoes from Qionghai city with MIR 7.1 (0-15.2) per 1,000. Oya virus (OYAV) and Bluetongue virus (BTV) were detected in biting midges from Wanning city with MIRs of 0.4 (0-1.2) and 0.1 (0-10.2) per 1,000, respectively. Three JEV strains were isolated and clustered within the genotype I group, which is presently the dominant genotype in China. Three TMUV strains were isolated for the first time on Hainan Island that belonged to Cluster 3. Three isolated GETVs were identified as Group 3. BTV was reported for the first time on Hainan Island, and the complete genome for one BTV strain was successfully assembled, which was classified as serotype 1 based on the sequences of segment 2. These results stress the need to develop adequate surveillance plan measures to better control the public health threat of arboviruses carried by mosquitoes and biting midges in local regions.

Cadmium is a hazardous heavy metal that is commonly found in the soil and poses significant risks to soil organisms. The toxic effects of Cd on soil microorganisms and earthworms (Eisenia foetida) have been extensively studied, but most studies focused on high Cd pollution levels. Therefore, this study aims to investigate the different responses of soil organisms to moderate and low levels of Cd contamination. According to the study, the presence of 2.5 mg/kg of Cd had a significant impact on the microbial community’s composition and diversity. The relative abundance of most microbes decreased, while the abundance of Firmicutes and Actinobacteriota showed a considerable increase. The LEFSE analysis revealed that the Bacillus genus of the Firmicutes phylum can serve as a biomarker in soil contaminated with 2.5 mg/kg of Cd. At the same time, the functional analysis of PICRUSt 2 shows that microorganisms found in polluted soil have a noticeable decrease in their ability to metabolize lipids. On the other hand, our findings indicate that Cd has a detrimental effect on the biomass of earthworms and induces oxidative stress in these animals. The activation of superoxide dismutase (SOD) and catalase (CAT) enzymes in earthworms was carried out to mitigate oxidative stress. The study found a strong positive relationship between SOD and both time and Cd pollution. However, CAT exhibited inhibition throughout the later stages of the experiment, particularly when exposed to relatively higher levels of pollution. The analysis of RNA in earthworms revealed that soil Cd pollution at a concentration of 2.5 mg/kg primarily impacts the cellular structure and function of earthworms. This pollution disrupts the integrity of the cytoskeleton structure, hampers DNA replication, and compromises the precision of cell signaling. Simultaneously, when compared to the control group, several metabolic pathways exhibited abnormalities.

At present, the application of "Internet +" in public health services is developing rapidly, but there is a lack of appropriate data standards, leading to difficulties in the construction process. In addition, the application of "Internet +" in public health services has both public welfare and commercial characteristics, involving multiple stakeholders and interested parties, resulting in imperfect development models, poor supply-demand matching, and insufficient sustainability. In this context, this paper selects two cases, the Shanghai Health Cloud Platform and the Vaccine Comprehensive Management Platform, to analyze the difficulties, solutions, and effectiveness of "Internet +" public health services. On this basis, the characteristics of "Internet+" public health applications are summarized, and corresponding countermeasures and suggestions are proposed to provide a reference for the theory and solutions of "Internet+" public health in other cities in China and global cities.

Abstract Background Mosquito-borne viruses cause various infectious diseases in humans and animals. Oya virus (OYAV) and Ebinur Lake virus (EBIV), belonging to the genus Orthobunyavirus within the family Peribunyaviridae, are recognized as neglected viruses with the potential to pose threats to animal or public health. The evaluation of vector competence is essential for predicting the arbovirus transmission risk. Methods To investigate the range of mosquito vectors for OYAV (strain SZC50) and EBIV (strain Cu20-XJ), the susceptibility of four mosquito species (Culex pipiens pallens, Cx. quinquefasciatus, Aedes albopictus, and Ae. aegypti) was measured through artificial oral infection. Then, mosquito species with a high infection rate (IR) were chosen to further evaluate the dissemination rate (DR), transmission rate (TR), and transmission efficiency. The viral RNA in each mosquito sample was determined by RT-qPCR. Results The results revealed that for OYAV, Cx. pipiens pallens had the highest IR (up to 40.0%) among the four species, but the DR and TR were 4.8% and 0.0%, respectively. For EBIV, Cx. pipiens pallens and Cx. quinquefasciatus had higher IR compared to Ae. albopictus (1.7%). However, the EBIV RNA and infectious virus were detected in Cx. pipiens pallens, with a TR of up to 15.4% and a transmission efficiency of 3.3%. Conclusions The findings indicate that Cx. pipiens pallens was susceptible to OYAV but had an extremely low risk of transmitting the virus. Culex pipiens pallens and Cx. quinquefasciatus were susceptible to EBIV, and Cx. pipiens pallens had a higher transmission risk to EBIV than Cx. quinquefasciatus. Graphical Abstract

Abstract Banna virus (BAV) is the prototype Seadornavirus, a class of reoviruses for which there has been little structural study. Here, we report atomic cryo-EM structures of three states of BAV virions—surrounded by 120 spikes (full virions), 60 spikes (partial virions), or no spikes (cores). BAV cores are double-layered particles similar to the cores of other non-turreted reoviruses, except for an additional protein component in the outer capsid shell, VP10. VP10 was identified to be a cementing protein that plays a pivotal role in the assembly of BAV virions by directly interacting with VP2 (inner capsid), VP8 (outer capsid), and VP4 (spike). Viral spikes (VP4/VP9 heterohexamers) are situated on top of VP10 molecules in full or partial virions. Asymmetrical electrostatic interactions between VP10 monomers and VP4 trimers are disrupted by high pH treatment, which is thus a simple way to produce BAV cores. Low pH treatment of BAV virions removes only the flexible receptor binding protein VP9 and triggers significant conformational changes in the membrane penetration protein VP4. BAV virions adopt distinct spatial organization of their surface proteins compared with other well-studied reoviruses, suggesting that BAV may have a unique mechanism of penetration of cellular endomembranes.

Introduction: Pancreatic cancer (PC) is a particularly aggressive malignancy with limited therapeutic options. The search for innovative treatments has focused on traditional Chinese medicine, specifically epimedium. This research investigates epimedium’s active ingredients, potential targets, and underlying mechanisms in treating PC.Methods: High-performance liquid chromatography (HPLC) was used to quantify the active components of epimedium and HPLC-Q-TOF-MS was employed for qualitative identification. Potential targets of epimedium’s active ingredients were identified using the TCMSP, ETCM, CTD, and Swiss Target Prediction databases. Potential PC-related targets were sourced from DisGeNET, GeneCards, and OMIM databases. A Venn diagram was utilized to identify overlapping PC-related and epimedium targets. Core targets and pathways were elucidated through protein-protein interaction (PPI) network analysis, Gene Ontology (GO) assessments, and Reactome pathway enrichment analyses. Molecular docking techniques investigated interactions between active compounds and these targets. The expression and prognostic implications of target genes were evaluated using GEPIA2 and the Human Protein Atlas (HPA) databases. In vitro studies assessed the impact of epimedium extract (EPE) on Panc-1 cell viability, and Western blot analysis examined the expression levels of key targets.Results: Network pharmacological indicate that epimedium econtains active components such as baohuoside I, icariin, hyperoside, and epimedin B, which have potential therapeutic effects against PC. In vitro assays confirmed that EPE significantly reduced the viability of Panc-1 cells. Western blot analysis revealed a considerable decrease in the expression of key targets in EPE-treated cells, including AKT1, EGFR, p-EGFR, JUN, BCL2, IL6, and SRC. The R-HSA-1280215: Interleukin-4 and Interleukin-13 signaling pathways involving these genes were identified as potential therapeutic targets.Discussion:Epimedium holds promise as a candidate for treating PC. The modulation of interleukin-4 and interleukin-13 signaling pathways could be a pivotal mechanism by which epimedium impedes tumor development. Further research is warranted to validate these findings and explore the clinical applicability of epimedium in PC treatment.

Objectives: Metagenomic next-generation sequencing (mNGS) is a powerful tool for pathogen detection. The accuracy depends on both wet lab and dry lab procedures. The objective of our study was to assess the influence of read length and dataset size on pathogen detection. Methods: In this study, 43 clinical BALF samples, which tested positive via clinical mNGS and were consistent with the diagnosis, were subjected to re-sequencing on the Illumina NovaSeq 6000 platform. The raw re-sequencing data, consisting of 100 million (M) paired-end 150 bp (PE150) reads, were divided into simulated datasets with eight different data sizes (5 M, 10 M, 15 M, 20 M, 30 M, 50 M, 75 M, 100 M) and five different read lengths (single-end 50 bp (SE50), SE75, SE100, PE100, and PE150). Both Kraken2 and IDseq bioinformatics pipelines were employed to analyze the previously diagnosed pathogens in the simulated data. Detection of pathogens was based on read counts ranging from 1 to 10 and RPM values ranging from 0.2 to 2. Results: Our results revealed that increasing dataset sizes and read lengths can enhance the performance of mNGS in pathogen detection. However, a larger data sizes for mNGS require higher economic costs and longer turnaround time for data analysis. Our findings indicate 20 M reads being sufficient for SE75 mode to achieve high recall rates. Additionally, high nucleic acid loads in samples can lead to increased stability in pathogen detection efficiency, reducing the impact of sequencing strategies. The choice of bioinformatics pipelines had a significant impact on recall rates achieved in pathogen detection. Conclusions: Increasing dataset sizes and read lengths can enhance the performance of mNGS in pathogen detection but increase the economic and time costs of sequencing and data analysis. Currently, the 20 M reads in SE75 mode may be the best sequencing option.

Abstract Background Metagenomic next-generation sequencing (mNGS) could improve the diagnosed efficiency of pathogens in bloodstream infections or sepsis. Little is known about the clinical impact of mNGS test when used for the early diagnosis of suspected infections. Herein, our main objective was to assess the clinical efficacy of utilizing blood samples to perform mNGS for early diagnosis of suspected infections, as well as to evaluate its potential in guiding antimicrobial therapy decisions. Methods In this study, 212 adult hospitalized patients who underwent blood mNGS test in the early stage of suspected infections were enrolled. Diagnostic efficacy of mNGS test and blood culture was compared, and the clinical impact of mNGS on clinical care was analyzed. Results In our study, the total detection rate of blood mNGS was significantly higher than that of culture method (74.4% vs. 12.1%, P

Abstract Background The escalation of viruses over the past decade has highlighted the need to determine their respective hosts, particularly for emerging ones that pose a potential menace to the welfare of both human and animal life. Yet, the traditional means of ascertaining the host range of viruses, which involves field surveillance and laboratory experiments, is a laborious and demanding undertaking. A computational tool with the capability to reliably predict host ranges for novel viruses can provide timely responses in the prevention and control of emerging infectious diseases. The intricate nature of viral-host prediction involves issues such as data imbalance and deficiency. Therefore, developing highly accurate computational tools capable of predicting virus-host associations is a challenging and pressing demand. Results To overcome the challenges of virus-host prediction, we present HostNet, a deep learning framework that utilizes a Transformer-CNN-BiGRU architecture and two enhanced sequence representation modules. The first module, k-mer to vector, pre-trains a background vector representation of k-mers from a broad range of virus sequences to address the issue of data deficiency. The second module, an adaptive sliding window, truncates virus sequences of various lengths to create a uniform number of informative and distinct samples for each sequence to address the issue of data imbalance. We assess HostNet's performance on a benchmark dataset of “Rabies lyssavirus” and an in-house dataset of “Flavivirus”. Our results show that HostNet surpasses the state-of-the-art deep learning-based method in host-prediction accuracies and F1 score. The enhanced sequence representation modules, significantly improve HostNet's training generalization, performance in challenging classes, and stability. Conclusion HostNet is a promising framework for predicting virus hosts from genomic sequences, addressing challenges posed by sparse and varying-length virus sequence data. Our results demonstrate its potential as a valuable tool for virus-host prediction in various biological contexts. Virus-host prediction based on genomic sequences using deep neural networks is a promising approach to identifying their potential hosts accurately and efficiently, with significant impacts on public health, disease prevention, and vaccine development.

Abstract Background Recent state-of-the-art sequencing technologies enable the investigation of challenging regions in the human genome and expand the scope of variant benchmarking datasets. Herein, we sequence a Chinese Quartet, comprising two monozygotic twin daughters and their biological parents, using four short and long sequencing platforms (Illumina, BGI, PacBio, and Oxford Nanopore Technology). Results The long reads from the monozygotic twin daughters are phased into paternal and maternal haplotypes using the parent–child genetic map and for each haplotype. We also use long reads to generate haplotype-resolved whole-genome assemblies with completeness and continuity exceeding that of GRCh38. Using this Quartet, we comprehensively catalogue the human variant landscape, generating a dataset of 3,962,453 SNVs, 886,648 indels (

Odontogenic stem cells originate from cranial neural crest cells and offer unique advantages in the regeneration of dentin-pulp complex. There is increasing evidence that stem cells exert their biological functions mainly through exosome-based paracrine effects. Exosomes contain DNA, RNA, proteins, metabolites, etc., which can play a role in intercellular communication and have similar therapeutic potential to stem cells. In addition, compared with stem cells, exosomes also have the advantages of good biocompatibility, high drug carrying capacity, easy to obtain, and few side effects. Odontogenic stem cell-derived exosomes mainly affect the regeneration of the dentin-pulp complex by regulating processes such as dentintogenesis, angiogenesis, neuroprotection and immunomodulation. This review aimed to describe “cell-free therapies” based on odontogenic stem cell-derived exosomes, which aim to regenerate the dentin-pulp complex.

ABSTRACTMosquito-borne viruses cause various infectious diseases in humans and animals. Tibet orbivirus (TIBOV), a newly identified arbovirus, efficiently replicates in different types of vertebrate and mosquito cells, with its neutralizing antibodies detected in cattle and goats. However, despite being isolated from Culicoides midges, Anopheles, and Culex mosquitoes, there has been a notable absence of systematic studies on its vector competence. Thus, in this study, Aedes aegypti and Culex pipiens pallens were reared in the laboratory to measure vector susceptibility through blood-feeding infection. Furthermore, RNA sequencing was used to examine the overall alterations in the Ae. aegypti transcriptome following TIBOV infection. The results revealed that Ae. aegypti exhibited a high susceptibility to TIBOV compared to Cx. p. pallens. Effective replication of the virus in Ae. aegypti midguts occurred when the blood-feeding titer of TIBOV exceeded 105 plaque-forming units mL−1. Nevertheless, only a few TIBOV RNA-positive samples were detected in the saliva of Ae. aegypti and Cx. p. pallens, suggesting that these mosquito species may not be the primary vectors for TIBOV. Moreover, at 2 dpi of TIBOV, numerous antimicrobial peptides downstream of the Toll and Imd signaling pathways were significantly downregulated in Ae. aegypti, indicating that TIBOV suppressed mosquitos’ defense to survive in the vector at an early stage. Subsequently, the stress-activated protein kinase JNK, a crucial component of the MAPK signaling pathway, exhibited significant upregulation. Certain genes were also enriched in the MAPK signaling pathway in TIBOV-infected Ae. aegypti at 7 dpi.IMPORTANCETibet orbivirus (TIBOV) is an understudied arbovirus of the genus Orbivirus. Our study is the first-ever attempt to assess the vector susceptibility of this virus in two important mosquito vectors, Aedes aegypti and Culex pipiens pallens. Additionally, we present transcriptome data detailing the interaction between TIBOV and the immune system of Ae. aegypti, which expands the knowledge about orbivirus infection and its interaction with mosquitoes.

Non-small cell lung cancer (NSCLC) is one of the most common cancers worldwide, and chemotherapy is one of its main treatment methods. However, there are significant differences in patients’ reactions to chemotherapy, leading to unsatisfactory treatment outcomes. Therefore, identifying relevant factors that affect the efficacy of chemotherapy can help doctors better develop personalized treatment plans, improve the treatment effectiveness, and quality of life of patients. This article aims to understand the specific clinical role of CYP1B1 gene in NSCLC. Therefore, based on the individualized health model of CYP1B1 gene polymorphism, this article analyzes the prediction of postoperative chemotherapy efficacy for NSCLC. Through a study on the control variables of postoperative recovery of stage III NSCLC in a hospital, according to the findings of this study, 14 of the 32 patients in the EGFR mutation-positive group relapsed. In the EGFR-negative group, 13 of the 36 patients relapsed. It can be considered that CYP1B1 gene polymorphism has a good curative effect in postoperative chemotherapy of NSCLC, and it can effectively control the recurrence rate of cancer.

Lung cancer is the cancer with the highest mortality rate and the highest incidence in the world at this stage. Among them, non-small lung cancer is the most common type of lung cancer, and most small cancers have disappeared, which is the optimal time for surgery at the time of diagnosis. To explore and systematically evaluate the clinical efficacy of CYP1B1 gene polymorphism in the treatment of epidermal growth factor receptor (EGFR) Mutant non-small cell lung cancer, this article proposes the principles of lung cancer screening based on CYP1B1 gene polymorphism and polarization imaging and explores the diagnosis and treatment of non-EGFR mutant lung cancer. Based on a large number of medical image data, imageomics can directly reflect the correlation between tumor molecular phenotype and image characteristics by deeply mining some imaging features of the image, which has important value in the early diagnosis of disease, the formulation of personalized treatment plan, and efficacy evaluation and prognosis prediction. A total of 141 NSCLC patients with sensitive EGFR mutation were included in this study, including 101 patients with EGFR single-gene mutation and 40 patients with EGFR multigene mutation coexisting mutation. Both groups of patients were female, aged ≥60 years, no smoking history, no family history of leukemia, adenocarcinoma, lung cancer, stage IV, lymph node metastasis, living, far from metastasis, and ECOG score of 0–2. This study examined the relative number of gene expression and PFS in EGFR multigene co-existing mutations. When the number of mixed genes is 1, 2, and higher, the PFS is 9 months, 8 months, and 6 months, respectively. The PFS time of this group of patients gradually shortened. Therefore, this study examined the benefit of polygenic mutation in estimation by comparing the clinical characteristics of patients with EGFR single-gene mutation and polygenic mutation, to provide measurement of EGFR-TKI and to provide suggestions for future drug selection.

Abstract Background New oral anticoagulants (NOACs) have been becoming prevalent in recent years and are increasingly used in the treatment of port vein thrombosis. The difference of the efficacy and safety between rivaroxaban and dabigatran remains unclear in the treatment of cirrhotic patients with acute portal vein thrombosis (PVT). Methods This retrospective study included all consecutive cirrhotic patients with acute portal vein thrombosis in our institute from January 2020 to December 2021. The patients received oral anticoagulation with rivaroxaban or dabigatran. The demographic, clinical, and imaging data of patients were collected. The diagnosis of acute PVT was confirmed by imaging examinations. The severity of liver cirrhosis was assessed using Child–Pugh score and Model for End-Stage Liver Disease (MELD) score. Outcomes included recanalization (complete, partial, and persistent occlusion), liver function, bleedings, and survival. The log-rank test was used to compare Kaplan–Meier distributions of time-to-event outcomes. The Cox proportional hazards model was used to calculate hazard ratios (HRs) with 95% confidence intervals (CIs). Results A total of 94 patients were included, 52 patients (55%) received rivaroxaban and 42 (45%) with dabigatran. The complete and partial recanalization of PVT was observed in 41 patients. There was no significant difference in complete recanalization, partial recanalization, and persistent occlusion between the two groups. With multivariate analysis, D-dimer (HR 1.165, 95% CI 1.036–1.311, p = 0.011) was independent predictors of complete recanalization. The Child–Pugh score (p = 0.001) was significantly improved in both two groups after anticoagulation, respectively. However, there was no difference between the two groups. The probability of survival was 94%, 95% in the rivaroxaban and dabigatran groups (log-rank p = 0.830). Major bleedings were reported in 3 patients (6%) in rivaroxaban group and 1 patient (2%) in dabigatran group (p = 0.646). Six patients (12%) in rivaroxaban group experienced minor bleeding, and five (12%) from dabigatran group (p = 0.691). Conclusions The efficacy and safety were comparable between rivaroxaban and dabigatran in the treatment of cirrhotic patients with acute portal vein thrombosis. And D-dimer can contribute to the prediction of PVT recanalization in cirrhotic patients.

Abstract Background Metagenomic next-generation sequencing (mNGS) is a novel nucleic acid method for the detection of unknown and difficult pathogenic microorganisms, and its application in the etiological diagnosis of fever of unknown origin (FUO) is less reported. We aimed to comprehensively assess the value of mNGS in the etiologic diagnosis of FUO by the pathogen spectrum and diagnostic performance, and to investigate whether it is different in the time to diagnosis, length of hospitalization, antibiotic consumption and cost between FUO patients with and without early application of mNGS. Methods A total of 149 FUO inpatients underwent both mNGS and routine pathogen detection was retrospectively analyzed. The diagnostic performance of mNGS, culture and CMTs for the final clinical diagnosis was evaluated by using sensitivity, specificity, positive predictive value, negative predictive value and total conforming rate. Patients were furtherly divided into two groups: the earlier mNGS detection group (sampling time: 0 to 3 days of the admission) and the later mNGS detection group (sampling time: after 3 days of the admission). The length of hospital stay, time spent on diagnosis, cost and consumption of antibiotics were compared between the two groups. Results Compared with the conventional microbiological methods, mNGS detected much more species and had the higher negative predictive (67.6%) and total conforming rate (65.1%). Patients with mNGS sampled earlier had a significantly shorter time to diagnosis (6.05+/-6.23 vs. 10.5+/-6.4 days, P

ABSTRACTAcinetobacter baumannii is a major opportunistic pathogen causing hospital-acquired infections, and it is imperative to comprehend its evolutionary and epidemiological dynamics in hospitals to prevent and control nosocomial transmission. Here, we present a comprehensive genomic epidemiological study involving the genomic sequencing and antibiotic resistance profiling of 634 A. baumannii strains isolated from seven intensive care units (ICUs) of a Chinese general hospital over 2 consecutive years. Our study reveals that ST2 is highly dominant (90.54%) in the ICUs, with 98.90% of the ST2 exhibiting multidrug resistant or extensively drug resistant. Phylogenetic analyses of newly sequenced genomes and public data suggest that nosocomial isolates originated outside the hospital but evolved inside. The major lineages appear to be stable, with 9 of the 28 identified nosocomial epidemic clones infecting over 60% of the affected patients. However, outbreaks of two highly evolved clones have been observed in different hospitals, suggesting significant inter-hospital transmission chains. By coupling patient medical records and genomic divergence of the ST2, we found that cross-ward patient transfer played a crucial role in pathogen’s nosocomial transmission. Additionally, we identified 831 potential adaptive evolutionary loci and 44 associated genes by grouping and comparing the genomes of clones with different prevalence. Overall, our study provides a comprehensive and contemporary survey on the epidemiology and genomic evolution of A. baumannii in a large Chinese general hospital. These findings shed light on the nosocomial evolution and transmission of A. baumannii and offers valuable information for transmission prevention and antibiotic therapy.IMPORTANCEThis study delved into the genomic evolution and transmission of nosocomial Acinetobacter baumannii on a large scale, spanning both an extended time period and the largest sample size to date. Through molecular epidemiological investigations based on genomics, we can directly trace the origin of the pathogen, detecting and monitoring outbreaks of infectious diseases in a timely manner, and ensuring public health safety. In addition, this study also collects a large amount of genomic and antibiotic resistance detection data, which is helpful for phenotype prediction based on genomic sequencing. It enables patients to receive personalized antibiotic treatment quickly, helps doctors select antibiotics more accurately, and contributes to reducing the use of antibiotics and lowering the risk of antibiotic resistance development.

Objectives: Oya virus (OYAV) and Ebinur lake virus (EBIV) belong to the genus Orthobunyavirus within the Peribunyaviridae family, and both are recognized as the novel virus with potential threat to the animal or public health. Given their potential to cause outbreaks and their detection in diverse samples across different regions, the need for a reliable and efficient molecular detection method for OYAV and EBIV becomes imperative. Methods: The S-segment of OYAV and EBIV was used for designing specific primer and probe sets, which were employed in a real-time reverse transcription quantitative PCR (RT-qPCR) assay. The analytical performance of these assays, encompassing specificity, sensitivity, reproducibility, and fitness for purpose, was thoroughly evaluated across various sample matrices. Results: The developed RT-qPCR assays were very specific to their respective targets. Both assays were highly reproducible (%CV

ABSTRACTPrevious studies have shown that the increased prevalent ST764 clone in China, Japan, and other Asian areas. However, the knowledge of the genetic features and virulence characteristics of methicillin-resistant Staphylococcus aureus (MRSA) ST764 in China is still limited. In this study, we identified 52 ST764-SCCmec type II isolates collected from five cities in China between 2014 and 2021. Whole genome sequencing showed that the most common staphylococcal protein A (spa) types of ST764 in China were t002 (55.78%) and t1084 (40.38%). Virulence assays showed that ST764-t1084 isolates had high haemolytic activity and α-toxin levels. Of the critical regulatory factors affecting α-toxin production, only the SaeRS was highly expressed in ST764-t1084 isolates. Mouse abscess model indicated that the virulence of ST764-t1084 isolates was comparable to that of S. aureus USA300-LAC famous for its hypervirulence. Interestingly, ST764-t002 isolates exhibited stronger biofilm formation and cell adhesion capacities than ST764-t1084 isolates. This seems to explain why ST764-t002 subclone has become more prevalent in China in recent years. Phylogenetic analysis suggested that all ST764 isolates from China in Clade III were closely related to KUN1163 (an isolate from Japan). Notably, genomic analysis revealed that the 52 ST764 isolates did not carry arginine catabolic mobile element (ACME), which differed from ST764 isolates in Japan. Additionally, most ST764 isolates (69.23%) harboured an obvious deletion of approximately 5 kb in the SCCmec II cassette region compared to KUN1163. Our findings shed light on the potential global transmission and genotypic as well as phenotypic characteristics of ST764 lineage.

Abstract Arthropod-borne virus (arbovirus) and arthropod-specific virus (ASV) are viruses circulating amongst hematophagous arthropods that are broadly transmitted in ecological systems. Arbovirus may replicate in both vertebrates and invertebrates and some are known to be pathogenic to animals or humans. ASV only replicate in invertebrate arthropods yet they are basal to many types of arboviruses. We built a comprehensive dataset of arbovirus and ASV by curating globally available data from the Arbovirus Catalog, the arbovirus list in Section VIII-F of the Biosafety in Microbiological and Biomedical Laboratories 6th edition, Virus Metadata Resource of International Committee on Taxonomy of Viruses, and GenBank. Revealing the diversity, distribution and biosafety recommendation of arbovirus and ASV at a global scale is essential to the understanding of potential interactions, evolution, and risks associated with these viruses. Moreover, the genomic sequences associated with the dataset will enable the investigation of genetic patterns distinguishing the two groups, as well as aid in predicting the vector/host relationships of the newly discovered viruses.

Abstract Background Rhizopus delemar is an invasive fungal pathogen that can cause fatal mucormycosis in immunodeficient individuals. Encephalitis caused by R. delemar is rare and difficult to diagnose early. Clinical detection methods for R. delemar include blood fungal culture, direct microscopic examination, and histopathological examination, but the detection is often inadequate for clinical diagnosis and can easily lead to missed diagnosis with delayed treatment. Case presentation We report a case of a 47-year-old male with brainstem hemorrhage caused by encephalitis due to R. delemar. The patient had a history of hypertension, type 2 diabetes, and irregular medication. No pathogens were detected in cerebrospinal fluid (CSF) and nasopharyngeal secretion cultures. R. delemar was identified by metagenomic next-generation sequencing (mNGS) in CSF, and in combination with the patient’s clinical characteristics, encephalitis caused by R. delemar was diagnosed. Antibiotic treatment using amphotericin B liposome in combination with posaconazole was given immediately. However, due to progressive aggravation of the patient’s symptoms, he later died due to brainstem hemorrhage after giving up treatment. Conclusions mNGS technique is a potential approach for the early diagnosis of infections, which can help clinicians provide appropriate antibiotic treatments, thus reducing the mortality and disability rate of patients.

Abstract Background Bacillus cereus infections in immunocompetent patients are uncommon and mainly observed in fragile patients. It can cause lethal infections with multiple organ dysfunction syndrome (MODS). However, a patient presenting as venous sinus thrombosis and survival without sequela has not been reported. Case presentation A 20-year-old previously healthy male developed gastroenteritis after a meal, followed by fever, convulsions, and severe disturbance of consciousness. The patient had significant leukocytosis with a mildly elevated D-dimer, creatinine level, and respiratory failure. The CT(computed tomography) revealed fatal brain edema and subarachnoid hemorrhage. Previous blood culture in a local hospital revealed B. cereus, which was confirmed by mNGS(metagenomic next-generation sequencing) using blood and urine in our hospital. Accordingly, B. cereus sepsis with MODS were considered. Later, cerebral venous sinus thrombosis was proved. After anti-infection (linezolid 0.6 g, Q12h; and meropenem 1.0 g, Q8h), anti-coagulant (enoxaparin 6000U, Q12h), and other symptomatic treatments, the patient recovered completely without sequela at the 6-month follow-up. Conclusions This case suggests that in immunocompetent adults, there is still a risk of infection with B. cereus, causing severe MODS. Special attention should be paid to venous sinus thrombosis and subarachnoid hemorrhage in such cases, while, anti-coagulant is essential therapy.

Abstract Introduction Lung cancer is the most prevalent cancer with high mortality in China, and it is associated with the dysbiosis of the lung microbiome. This study attempted to screen for specific microorganisms as potential biomarkers for distinguishing benign lung disease from lung cancer. Methods Bronchoalveolar lavage fluid (BALF) sample was selected in the study instead of saliva to avoid contamination with oral microorganisms, and microbial taxonomic and functional differences in BALF samples from patients with lung cancer and those with those from patients with benign lung diseases were performed based on metagenomic next‐generation sequencing, for the first time, so that microorganisms other than bacteria could be included. Results The results showed that the intrasample diversity of malignant samples was different from benign samples, and the microbial differences among malignant samples were smaller, with lower microbial diversity, significantly changed microbial abundance and metabolic functions. Metabolic function analysis revealed amino acid‐related metabolism was more prevalent in benign samples, whereas carbohydrate‐related metabolism was more prevalent in malignant samples. By LEfSe, Metastat and Random Forest analysis, we identified a series of important differential microorganisms. Importantly, the model combining five key genera plus one tumor marker (neuron‐specific enolase) as indicators presented the optimal disease typing performance. Conclusion Thus results suggest the value of these differential microorganisms enriched in tumors in mechanism research and may be potential new targets for lung cancer therapy. More importantly, the biomarkers identified in this study can be conducive to improve the clinical diagnosis of lung cancer and have good application prospects.

Orthobunyavirus is the largest and most diverse genus in the family Peribunyaviridae. Orthobunyaviruses are widely distributed globally and pose threats to human and animal health. Ebinur Lake virus (EBIV) is a newly classified Orthobunyavirus detected in China, Russia, and Kenya. This study explored the antiviral effects of two broad-spectrum antiviral drugs, favipiravir and ribavirin, in a BALB/c mouse model. Favipiravir significantly improved the clinical symptoms of infected mice, reduced viral titer and RNA copies in serum, and extended overall survival. The median survival times of mice in the vehicle- and favipiravir-treated groups were 5 and 7 days, respectively. Favipiravir significantly reduced virus titers 10- to 100-fold in sera at all three time points compared to vehicle-treated mice. And favipiravir treatment effectively reduced the virus copies by approximately 10-fold across the three time points, relative to vehicle-treated mice. The findings expand the antiviral spectrum of favipiravir for orthobunyaviruses in vivo.

Abstract We present multiPrime, a novel tool that automatically designs minimal primer sets for targeted next‐generation sequencing, tailored to specific microbiomes or genes. MultiPrime enhances primer coverage by designing primers with mismatch tolerance and ensures both high compatibility and specificity. We evaluated the performance of multiPrime using a data set of 43,016 sequences from eight viruses. Our results demonstrated that multiPrime outperformed conventional tools, and the primer set designed by multiPrime successfully amplified the target amplicons. Furthermore, we expanded the application of multiPrime to 30 types of viruses and validated the work efficacy of multiPrime‐designed primers in 80 clinical specimens. The subsequent sequencing outcomes from these primers indicated a sensitivity of 94% and a specificity of 89%.

Dengue fever (DF) and dengue hemorrhagic fever (DHF) are among the most common tropical diseases affecting humans. To analyze the risk of clinical and transmission of DF/DHF in Shenzhen, the surveillance on patients of all-age patients with dengue virus (DENV) infections was conducted. Our findings revealed that the majority of DENV-infected patients are young to middle-aged males, and the development of the disease is accompanied by abnormal changes in the percentages of neutrophils, lymphocytes, and basophils. Demographic analysis revealed that these patients is concentrated in areas such as Futian District, which may be due to the higher mosquito density and temperature than that in other area. Subsequent, mosquito infection experiments confirmed that the effect of temperature shift on DENV proliferation and transmission. Not only that, constant temperatures can enhance the spread of DENV, even increase the risk of epidemic. Thus, the role of innate immune response should be highlighted in the prediction of severe severity of DENV-infected patients, and temperature should be taken into account in the prevention and control of DENV. Introduction: Dengue fever (DF) and dengue hemorrhagic fever (DHF) are among the most common tropical diseases affecting humans, and which caused by the four dengue virus serotypes (DENV 1–4). Objectives: To analyze the risk of clinical and transmission of DF/DHF in Shenzhen. Methods: The surveillance on patients of all-age patients with dengue virus (DENV) infections was conducted. Results: Our findings revealed that the majority of DENV-infected patients are young to middle-aged males, and the development of the disease is accompanied by abnormal changes in the percentages of neutrophils, lymphocytes, and basophils. Demographic analysis revealed that these patients is concentrated in areas such as Futian District, which may be due to the higher mosquito density and temperature than that in other area. Subsequent, mosquito infection experiments confirmed that the effect of temperature shift on DENV proliferation and transmission. Not only that, constant temperatures can enhance the spread of DENV, even increase the risk of epidemic. Conclusion: 1. Elevated levels of neutrophils, lymphocytes, basophils, and temperature are all significant risk factors for dengue transmission and pathogenesis; 2. Temperature increasing is associated with a higher risk of dengue transmission; 3. Fluctuations in temperature around 28 °C (28 ± 5 °C) would increase dengue transmission.

Abstract New and old subgrade stability is a crucial problem for widening projects in soft soil foundations because uncoordinated deformations can cause serious engineering accidents, even disasters. In order to ensure the stability of a widening expressway project near a pond in the Qinbei section, a series of on-site investigations, lab tests, and numerical analyses was performed. The settlement and displacement of on-site monitoring were carried out during the construction period to verify the analyzed results. It was found that the uneven settlement caused heavy settlement and displacement of the embankment; thus, net cracks and longitudinal cracks occurred in the expressway. The new embankment was also affected by the weak foundation; therefore, the foundation should be reinforced before the construction of the widening part. Considering the economy and effectiveness of the widening project, the replacement method was recommended for the weak foundation reinforcement based on the numerical analysis and on-site motoring results. Water pumping had a significant effect on the settlement of the embankment. Water pumping caused seepage in the foundation and increased the effective stress of the soil, making the foundation more consolidated. The pond slope should be reinforced before the construction of the embankment because it was not confined by the surrounding soil. Therefore, the slope soil could not provide sufficient passive soil pressure and easily slide.

The long noncoding RNA DARS-AS1 was aberrantly expressed and participated in several human cancer progressions, whereas whether DARS-AS1 is involved in human gastric cancer remains unclear. This study aimed to investigate the influence of DARS-AS1 on gastric cancer progression and explore the potential regulatory network of DARS-AS1/miR-330-3p/NAT10. The expression levels of DARS-AS1, miR-330-3p, and NAT10 were measured by quantitative real-time polymerase chain reaction. The CCK-8 assay and Transwell assay were used to determine the cell viability, migration, and invasion capacities, respectively. The target association between miR-330-3p and DARS-AS1 or NAT10 was confirmed using a luciferase reporter assay. In result, DARS-AS1 levels were elevated in tumor tissues and associated with shorter overall survival in patients with gastric cancer. Knockdown of DARS-AS1 could hamper cell viability, migration, and invasion in gastric cancer cells. DARS-AS1 acts as a competitive endogenous RNA to regulate the NAT10 expression by sponging miR-330-3p in gastric cancer cells. In conclusion, DARS-AS1 was elevated in gastric cancer, and DARS-AS1/miR-330-3p/NAT10 signaling offered some new horizons for predicting prognosis and a novel therapeutic method for the treatment of gastric cancer.

Biting midges are one of the most common hematophagous insects. They are capable of transmitting a wide range of arboviruses and have a significant impact on public health and veterinary medicine. Herein, from midge samples collected in 2013 in Yunnan, China, one sample induced a cell cytopathic effect (CPE) in BHK-21, MA104, and PK15 cell lines. Next-generation sequencing data, RACE and PCR determined the genome sequence of the sample and designated as an Oya virus (OYAV) isolate SZC50. Phylogenetic analysis of the sample revealed that it was cluster into viruses from species Orthobunyavirus catqueense. The open reading frames of S, M, and L segment of OYAV SZC50 were closest to those of OYAV SC0806. Moreover, 831 serum samples (736 pigs, 45 cattle, and 50 sheep) were gathered from 13 cities in Yunnan Province to detect neutralizing antibody of OYAV SZC50. A significant proportion of OYAV SZC50 antibody (more than 30%) was found in Yunnan pig populations, with the positive rate of OYAV SZC50 antibody in pigs from Malipo reaching 95%. To determine the pathogenicity of OYAV SZC50, we chose three animal models: specific pathogen-free Kunming mice, C57BL/6 mice lacking the interferon α/β receptor, and chicken embryos. At 5, 6, and 7 days post-infection, all adult and suckling C57BL/6 mice, and specific pathogen-free suckling Kunming mice were dead. Our finding was expanding the knowledge about the infection and pathogenic risk of the neglected virus in the Orthobunyavirus.

Abstract Background Burkholderia pseudomallei is a tropical pathogen that causes melioidosis. Its intrinsic drug-resistance is a leading cause of treatment failure, and the few available antibiotics require prolonged use to be effective. This study aimed to assess the clinical potential of B. pseudomallei phages isolated from Hainan, China. Methods Burkholderia pseudomallei strain (HNBP001) was used as the isolation host, and phages were recovered from domestic environmental sources, which were submitted to the host range determination, lytic property assays, and stability tests. The best candidate was examined via the transmission electron microscope for classification. With its genome sequenced and analyzed, its protective efficacy against B. pseudomallei infection in A549 cells and Caenorhabditis elegans was evaluated, in which cell viability and survival rates were compared using the one-way ANOVA method and the log-rank test. Results A phage able to lyse 24/25 clinical isolates was recovered. It was classified in the Podoviridae family and was found to be amenable to propagation. Under the optimal multiplicity of infection (MOI) of 0.1, an eclipse period of around 20 min and a high titer (1012 PFU/ml) produced within 1 h were demonstrated. This phage was found stabile at a wide range of temperatures (24, 37, 40, 50, and 60 °C) and pH values (3–12). After being designated as vB_BpP_HN01, it was fully sequenced, and the 71,398 bp linear genome, containing 93 open reading frames and a tRNA-Asn, displayed a low sequence similarity with known viruses. Additionally, protective effects of applications of vB_BpP_HN01 (MOI = 0.1 and MOI = 1) alone or in combination with antibiotics were found to improve viability of infected cells (70.6 ± 6.8%, 85.8 ± 5.7%, 91.9 ± 1.8%, and 96.8 ± 1.8%, respectively). A significantly reduced mortality (10%) and a decreased pathogen load were demonstrated in infected C. elegans following the addition of this phage. Conclusions As the first B. pseudomallei phage was isolated in Hainan, China, phage vB_BpP_HN01 was characterized by promising lytic property, stability, and efficiency of bacterial elimination during the in vitro/vivo experiments. Therefore, we can conclude that it is a potential alternative agent for combating melioidosis. Graphical Abstract

Human infection with Chlamydia psittaci is rare but difficult to recognize. We report an outbreak of psittacosis among poultry processing workers in China. We applied metagenomic next-generation sequencing and identified Chlamydia psittaci reads from samples of all seven patients, four of which were subsequently confirmed by PCR. Epidemiological results suggested that the poultry processed in the factory was the possible source of human infection.

Biological threats, whether naturally occurring or accidentally or deliberately released, have the potential to endanger lives and disrupt economies worldwide. Thus, high-containment protective measures should be implemented when handling bio-agents that can cause serious, highly contagious diseases, otherwise devastating pandemics can occur. In the fields of scientific investigation, healthcare, and product development, high-containment facilities play a critical role in preventing, detecting, and responding promptly and effectively to threats to global health security. In this paper, we present a summary of the main types of high-containment facilities, as well as their applications, challenges, and suggestions for the future.

Abstract Background The association between osteocalcin and mortality has been scantly studied. We aimed to investigate the association between osteocalcin along with its trajectories and mortality based on long-term longitudinal data. Methods We performed a retrospective cohort study of 9413 type 2 diabetic patients with at least three measurements of total serum osteocalcin within 3 years since their first inpatient diagnosis of type 2 diabetes. Baseline, mean values of osteocalcin levels and their trajectories were used as exposures. A multivariable-adjusted Cox proportional hazards model was used to estimate the association of osteocalcin levels and their trajectories with mortality. Results During a mean follow-up of 5.37 years, 1638 patients died, of whom 588 were due to cardiovascular events. Multivariable-adjusted hazard ratios (HRs) across quintiles of baseline osteocalcin levels were 2.88 (95% confidence interval (CI) 2.42–3.42), 1.65 (95% CI 1.37–1.99), 1.17 (95% CI 0.96–1.42), 1.00, and 1.92 (95% CI 1.60–2.30) for all-cause mortality, and 3.52 (95% CI 2.63–4.71), 2.00 (95% CI 1.46–2.73), 1.03 (95% CI 0.72–1.47), 1.00, 1.67 (95% CI 1.21–2.31) for CVD mortality, respectively. When we used the mean values of osteocalcin as the exposure, U-shaped associations were also found. These U-shaped associations were consistent among patients of different baseline characteristics. Patients with a stable or even increasing trajectory of osteocalcin may have a lower risk of both all-cause and CVD mortality. Conclusions A U-shape association between baseline osteocalcin and mortality was observed among patients with type 2 diabetes. Patients with lower levels of serum osteocalcin during follow-ups had higher risks for all-cause and cardiovascular mortality.

IntroductionPolymyxin-resistant Enterobacterales poses a significant threat to public health globally, but its prevalence and genomic diversity within a sole hospital is less well known. In this study, the prevalence of polymyxin-resistant Enterobacterales in a Chinese teaching hospital was investigated with deciphering of their genetic determinants of drug resistance.MethodsPolymyxin-resistant Enterobacterales isolates identified by matrix-assisted laser desorption were collected in Ruijin Hospital from May to December in 2021. Both the VITEK 2 Compact and broth dilution methods were used to determine polymyxin B (PMB) susceptibility. Polymyxin-resistant isolates were further characterized by molecular typing using PCR, multi-locus sequence typing, and sequencing of the whole genome.ResultsOf the 1,216 isolates collected, 32 (2.6%) across 12 wards were polymyxin-resistant (minimum inhibitory concentration (MIC) range, PMB 4–256 mg/ml, and colistin 4 ≥ 16 mg/ ml). A total of 28 (87.5%) of the polymyxin-resistant isolates had reduced susceptibility to imipenem and meropenem (MIC ≥ 16 mg/ml). Of the 32 patients, 15 patients received PMB treatment and 20 survived before discharge. The phylogenetic tree of these isolates showed they belonged to different clones and had multiple origins. The polymyxin-resistant Klebsiella pneumoniae isolates belonged to ST-11 (85.72%), ST-15 (10.71%), and ST-65 (3.57%), and the polymyxin-resistant Escherichia coli belonged to four different sequence types, namely, ST-69 (25.00%), ST-38 (25.00%), ST-648 (25.00%), and ST-1193 (25.00%). In addition, six mgrB specific mutations (snp_ALT c.323T>C and amino acid change p.Val8Ala) were identified in 15.6% (5/32) of the isolates. mcr-1, a plasmid-mediated polymyxin-resistant gene, was found in three isolates, and non-synonymous mutations including T157P, A246T, G53V, and I44L were also observed.DiscussionIn our study, a low prevalence of polymyxin-resistant Enterobacterales was observed, but these isolates were also identified as multidrug resistant. Therefore, efficient infection control measures should be implemented to prevent the further spread of resistance to last-line polymyxin therapy.

BackgroundMetagenomic next-generation sequencing (mNGS) is a promising technology that allows unbiased pathogen detection and is increasingly being used for clinical diagnoses. However, its application in urinary tract infection (UTI) is still scarce.MethodsThe medical records of 33 patients with suspected UTI who were admitted to the Second Hospital of Tianjin Medical University from March 2021 to July 2022 and received urine mNGS were retrospectively analyzed. The performance of mNGS and conventional urine culture in diagnosing infection and identifying causative organisms was compared, and the treatment effects were evaluated in terms of changes in urinalyses and urinary symptoms.ResultsIn the detection of bacteria and fungi, mNGS detected at least one pathogen in 29 (87.9%) cases, including 19 (57.6%) with positive mNGS but negative culture results and 10 (30.3%) with both mNGS and culture positive results. The remaining 4 (12.1%) patients were negative by both tests. Overall, mNGS performed better than culture (87.9% vs. 30.3%, P < 0.001). Within the 10 double-positive patients, mNGS matched culture results exactly in 5 cases, partially in 4 cases, and not at all in 1 case. In addition, mNGS detected a broader pathogen spectrum, detecting 26 species compared to only 5 species found in culture. The most abundant bacteria detected by mNGS was Escherichia coli, detected in 9 (27.2%) patients. All anaerobic bacteria, Mycobacterium Tuberculosis and all mixed pathogens were detected by mNGS. The final clinical diagnosis of UTI was made in 25 cases, and the sensitivity of mNGS was significantly higher than culture (100.0% vs 40.0%; P < 0.001) when using the diagnosis as a reference standard; the positive predictive value, negative predictive value and specificity were 86.2%, 100% and 50.0%, respectively. Importantly, targeted antibiotic therapy based on mNGS resulted in significant improvement in urinalyses and urinary symptoms in patients.ConclusionsmNGS is a technology that has shown clear advantages over culture, particularly in the context of mixed infections and UTIs that are difficult to diagnose and treat. It helps to improve the detection of pathogens, guide changes in treatment strategies, and is an effective complement to urine culture.

Abstract Background Patients with allogeneic hematopoietic stem cell transplantation (allo-HSCT) are prone to complicate viral infection. Central nervous system (CNS) involvement caused by the viruses is rare but with poor prognosis. Hantavirus, which usually cause hemorrhagic fever with renal syndrome (HFRS), and none case has been reported about these infection in allo-HSCT patients. Case presentation In August 2021, a 13-year-old male child developed intermittent fever and refractory hypotension after allo-HSCT. Magnetic resonance imaging of the head revealed abnormal signal foci in the left midbrain cerebral peduncle and bilateral thalamus. His family reported traces of mouse activity in the patient’s home kitchen. HFRS was suspected, but with no significant kidney damage. The specific immunoglobulin (Ig) G and M of hantavirus were negative. The metagenomic next-generation sequencing (mNGS) detected Seoul Orthohantavirus (SEOV) sequences directly in cerebrospinal fluid and blood. Conclusions Allo-HSCT patients are a high-risk group for infection. Usually the causative agent of infection is difficult to determine, and sometimes the site of infection is concealed. This report highlights the importance of suspecting hantavirus infection in allo-HSCT patients with CNS symptoms despite the absence of renal syndromes. The mNGS is a powerful tool for detecting pathogens. CNS infection with Seoul orthohantavirus in transplant patients is rare but possible as demonstrated in this case. To the best of our knowledge, this is the first reported case employing mNGS to diagnose SEOV caused CNS infection in an allo-HSCT patient.

Objective: Infection is one of the most common causes of death in children with hematological diseases. Here, we aim to investigate the value of metagenomic next-generation sequencing (mNGS) in the detection of causative pathogens in children with hematological diseases. Methods: In this retrospective study, specimens from children with hematological diseases, who were admitted to Sun Yat-Sen University between June 2019 and September 2021, were collected for culture and mNGS. Results: A total of 67 pediatric patients were enrolled, and 96 specimens were collected. The positive rate of mNGS was significantly higher than that of culture (57.2% vs 12.5%, P

ObjectiveTo investigate the diagnostic value of metagenomic next-generation sequencing (mNGS) using pleural effusion and ascites from children with sepsis.MethodsIn this study, children with sepsis or severe sepsis and appeared pleural or peritoneal effusions were enrolled, of whom the pleural effusions or ascites and blood samples were conducted pathogen detection using both conventional and mNGS methods. The samples were divided into pathogen-consistent and pathogen-inconsistent groups based on the consistency of mNGS results from different sample types, and into exudate and transudate groups based on their pleural effusion and ascites properties. The pathogen positive rates, pathogen spectrum, consistency between different sample types, and clinical diagnosis consistency were compared between mNGS and conventional pathogen tests.ResultsA total of 42 pleural effusions or ascites and 50 other type samples were collected from 32 children. The pathogen positive rate of the mNGS test was significantly higher than that of traditional methods (78.57% vs. 14.29%, P < 0.001) in pleural effusion and ascites samples, with a consistent rate of 66.67% between the two methods. Nearly 78.79% (26/33) of mNGS positive results of the pleural effusions and ascites samples were consistent with clinical evaluation, and 81.82% (27/33) of these positive samples reported 1-3 pathogens. The pathogen-consistent group outperformed the pathogen-inconsistent group in terms of consistency with respect to clinical evaluation (88.46% vs. 57.14%, P = 0.093), while there was no significant difference between the exudate and transudate groups (66.67% vs. 50.00%, P = 0.483).ConclusionCompared to conventional methods, mNGS has great advantages in pathogen detection of pleural effusion and ascites samples. Moreover, consistent results of mNGS tests with different sample types provide more reference values in clinical diagnosis.

ABSTRACT Central nervous system (CNS) infections can cause significant morbidity and mortality, especially in children. Rapid and accurate pathogenic detection in suspected CNS infections is essential for disease control at the early stage of infection. To evaluate the performance of metagenomic next-generation sequencing (mNGS) of cell-free DNA (cfDNA) in cerebrospinal fluid (CSF) in pediatric patients, we retrospectively analyzed the efficiency of cfDNA mNGS in children with CNS infections (n = 257) or noninfectious neurological disorders (n = 81). The CSF samples of 124 random subjects were used to evaluate the accuracy between mNGS of cfDNA and whole-cell DNA (wcDNA). In total, cfDNA mNGS detected a wide range of microbes with a detection rate of 71.0%, and the sensitivity and total coincidence rate with clinical diagnosis reached 68.9% and 67.5%, respectively. Compared with wcDNA mNGS, cfDNA mNGS had a higher efficacy in detecting viruses (66 versus 13) and Mycobacterium (7 versus 1), with significantly higher reads per million. The dominant causative pathogens were bacteria and viruses in CNS infections, but these presented with different pathogen spectra in different age categories. The best timing for the mNGS test ranged from 1 to 6 days after the start of anti-infection therapy, and the earlier mNGS started, the better was identification of bacterial CNS infections. This study emphasized that cfDNA mNGS had overall superiority to conventional methods on causative pathogen detection in pediatric CNS infections, and it was even better than wcDNA mNGS. Furthermore, research needs to be better validated in large-scale clinical trials to improve the clinical applications of cfDNA mNGS. IMPORTANCE Our study emphasized that cfDNA mNGS had overall superiority to conventional methods on causative pathogen detection in CNS-infected children, and it was even better than wcDNA mNGS. cfDNA mNGS detected a wide range of pathogens with a high total coincidence rate (67.5%) against clinical diagnosis. The best timing for cfDNA mNGS detection ranged from 1 to 6 days, rather than 0 days, after the start of empirical anti-infection therapy. The earlier mNGS started, the better the identifications of bacterial CNS infections. To the best of our knowledge, this research is the first report evaluating the clinical utility of mNGS with different methods (cfDNA versus wcDNA) of extracting DNA from CSF specimens in diagnosing pediatric CNS infections. Meanwhile, this is the largest cohort study that has evaluated the performance of mNGS using cfDNA from CSF specimens in pediatric patients with CNS infections.

ABSTRACT Infectious diseases pose a fatal risk to neonates. Timely and accurate pathogen detection is crucial for proper clinical diagnosis and therapeutic strategies. Limited sample volumes from neonatal patients seriously hindered the accurate detection of pathogens. Here, we unravel that metagenomic next-generation sequencing (mNGS) of cell-free DNA (cfDNA) and RNA can achieve unbiased detection of trace pathogens from different kinds of body fluid samples and blood samples. We enrolled 168 neonatal patients with suspected infections from whom blood samples (n = 153), cerebrospinal fluid samples (n = 127), and respiratory tract samples (RTSs) (including bronchoalveolar lavage fluids, sputa, and respiratory secretions) (n = 51) were collected and analyzed using mNGS. High rates of positivity (70.2%; 118/168) of mNGS were observed, and the coincidence rate against the final clinical diagnosis in positive mNGS cases reached 68.6% (81/118). The most common causative pathogens were Klebsiella pneumoniae (n = 12), Escherichia coli (n = 12), and Streptococcus pneumoniae (n = 8). mNGS using cfDNA and RNA can identify microbes that cannot be detected by conventional methods in different body fluid and blood samples, and more than 50% of these microbes were identified as causative pathogens. Further local polynomial regression fitting analysis revealed that the best timing for mNGS detection ranged from 1 to 3 days after the start of continuous antimicrobial therapy. Diagnosed and guided by mNGS results, the therapeutic regimens for 86 out of 117 neonatal patients were changed, most of whom (80/86) completely recovered and were discharged, while 44 out of 86 patients completely or partially stopped unnecessary medication. Our findings highlight the importance of mNGS in detecting causative DNA and RNA pathogens in infected neonatal patients. IMPORTANCE To the best of our knowledge, this is the first report on evaluating the performance of mNGS using cfDNA and RNA from body fluid and blood samples for diagnosing neonatal infections. mNGS of RNA and cfDNA can achieve the unbiased detection and identification of trace pathogens from different kinds of neonatal body fluid and blood samples with a high total coincidence rate (226/331; 68.3%) against final clinical diagnoses by sample. The best timing for mNGS detection in neonatal infections ranged from 1 to 3 days, rather than 0 days, after the start of continuous antimicrobial therapy. Our findings highlight the importance of mNGS in detecting causative DNA and RNA pathogens, and the extensive application of mNGS for the diagnosis of neonatal infections can be expected.

Abstract Large-scale COVID-19 vaccinations are currently underway in many countries in response to the COVID-19 pandemic. Here, we report, besides generation of neutralizing antibodies, consistent alterations in hemoglobin A1c, serum sodium and potassium levels, coagulation profiles, and renal functions in healthy volunteers after vaccination with an inactivated SARS-CoV-2 vaccine. Similar changes had also been reported in COVID-19 patients, suggesting that vaccination mimicked an infection. Single-cell mRNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) before and 28 days after the first inoculation also revealed consistent alterations in gene expression of many different immune cell types. Reduction of CD8+ T cells and increase in classic monocyte contents were exemplary. Moreover, scRNA-seq revealed increased NF-κB signaling and reduced type I interferon responses, which were confirmed by biological assays and also had been reported to occur after SARS-CoV-2 infection with aggravating symptoms. Altogether, our study recommends additional caution when vaccinating people with pre-existing clinical conditions, including diabetes, electrolyte imbalances, renal dysfunction, and coagulation disorders.

Although the fast-growing metagenomic next-generation sequencing (mNGS) has been used in diagnosing infectious diseases, low detection rate of mNGS in detecting pathogens with low loads limits its extensive application. In this study, 130 patients with suspected pulmonary infections were enrolled, from whom bronchoalveolar lavage fluid (BALF) samples were collected. The conventional tests and mNGS of cell-free DNA (cfDNA) and whole-cell DNA (wcDNA) using BALF were simultaneously performed. mNGS of cfDNA showed higher detection rate (91.5%) and total coincidence rate (73.8%) than mNGS of wcDNA (83.1% and 63.9%) and conventional methods (26.9% and 30.8%). A total of 70 microbes were detected by mNGS of cfDNA, and most of them (60) were also identified by mNGS of wcDNA. The 31.8% (21/66) of fungi, 38.6% (27/70) of viruses, and 26.7% (8/30) of intracellular microbes can be only detected by mNGS of cfDNA, much higher than those [19.7% (13/66), 14.3% (10/70), and 6.7% (2/30)] only detected by mNGS of wcDNA. After in-depth analysis on these microbes with low loads set by reads per million (RPM), we found that more RPM and fungi/viruses/intracellular microbes were detected by mNGS of cfDNA than by mNGS of wcDNA. Besides, the abilities of mNGS using both cfDNA and wcDNA to detect microbes with high loads were similar. We highlighted the advantage of mNGS using cfDNA in detecting fungi, viruses, and intracellular microbes with low loads, and suggested that mNGS of cfDNA could be considered as the first choice for diagnosing pulmonary infections.

BackgroundCentral nervous system (CNS) infection is one of the most serious complications after neurosurgery. Traditional clinical methods are difficult to diagnose the pathogen of intracranial infection. Due to recent advances in genomic approaches, especially sequencing technologies, metagenomic next-generation sequencing (mNGS) has been applied in many research and clinical settings.Case presentationHere, we report a case of CNS infection with Acinetobacter baumannii in a 15-year-old woman, who previously underwent surgery for recurrence of ependymoma in the fourth ventricle. On the eleventh postoperative day, the patient had a high fever and leukocytosis in the cerebrospinal fluid (CSF). mNGS using CSF rapidly and accurately identified the causative pathogen as A. baumannii with carbapenem resistance genes blaOXA-23 and blaOXA-51, which were confirmed by subsequent culture and susceptibility tests within 5 days. During the disease, mNGS, culture, and drug susceptibility testing were continued to monitor changes in pathogenic bacteria and adjust medication. At present, there are no case reports on to the use of mNGS for detecting pathogens in postoperative infection with ependymoma and guide medication.ConclusionmNGS has great advantages in pathogen identification and even pathogen resistance prediction. Multiple mNGS examinations during the course of the disease play an important role in the dynamic monitoring of pathogens.