A Drug Molecule-Modified Graphene Field-Effect Transistor Nanosensor for Rapid, Label-Free, and Ultrasensitive Detection of Estrogen Receptor α Protein.

Estrogen receptor α (ERα) is an important biomarker in breast cancer diagnosis and treatment. Sensitive and accurate detection of ERα protein expression is crucial in guiding selection of an appropriate therapeutic strategy to improve the effectiveness and prognosis of breast cancer treatment. Herein, we report a liquid-gated graphene field-effect transistor (FET) biosensor that enables rapid, sensitive, and label-free detection of the ERα protein by employing a novel drug molecule as a capture probe. The drug molecule was synthesized and subsequently immobilized onto the sensing surface of the fabricated graphene FET, which was able to distinguish the ERα-positive from the ERα-negative protein. The developed sensor not only demonstrated a low detection limit (LOD: 2.62 fM) but also achieved a fast response to ERα protein samples within 30 min. Moreover, depending on the relationship between the change of dirac point and the ERα protein concentrations, the dissociation constant (Kd) was estimated to be 7.35 ± 0.06 pM, indicating that the drug probe-modified graphene FET had a good affinity with ERα protein. The nanosensor was able to analyze ERα proteins from 36 cell samples lysates. These results show that the graphene FET sensor was able to differentiate between ERα-positive and ERα-negative cells, indicating a promising biosensor for the ultrasensitive and rapid detection of ERα protein without antibody labeling.

A portable transistor immunosensor for fast identification of porcine epidemic diarrhea virus.

Widespread distribution of porcine epidemic diarrhea virus (PEDV) has led to catastrophic losses to the global pig farming industry. As a result, there is an urgent need for rapid, sensitive and accurate tests for PEDV to enable timely and effective interventions. In the present study, we develop and validate a floating gate carbon nanotubes field-effect transistor (FG CNT-FET)-based portable immunosensor for rapid identification of PEDV in a sensitive and accurate manner. To improve the affinity, a unique PEDV spike protein-specific monoclonal antibody is prepared by purification, and subsequently modified on FG CNT-FET sensor to recognize PEDV. The developed FET biosensor enables highly sensitive detection (LoD: 8.1 fg/mL and 100.14 TCID50/mL for recombinant spike proteins and PEDV, respectively), as well as satisfactory specificity. Notably, an integrated portable platform consisting of a pluggable FG CNT-FET chip and a portable device can discriminate PEDV positive from negative samples and even identify PEDV and porcine deltacoronavirus within 1 min with 100% accuracy. The portable sensing platform offers the capability to quickly, sensitively and accurately identify PEDV, which further points to a possibility of point of care (POC) applications of large-scale surveillance in pig breeding facilities.

Coexistence of Superhardness and Metal-Like Electrical Conductivity in High-Entropy Dodecaboride Composite with Atomic-Scale Interlocks.

High electrical conductivity and super high hardness are two sought-after material properties, but both are contradictory because the effective suppression of dislocation movement generally increases the scattering of conducting electrons. Here we synthesized a high-entropy dodecaboride composite (HEDC) with a large number of atomic-scale interlocking layers. It shows a Vickers hardness of 51.2 ± 3.6 GPa under an applied load of 0.49 N and an electrical resistivity of 44.5 µΩ·cm at room temperature. Such HEDC achieves superhardness by inheriting the high intrinsic hardness of its constituent phases and restricting the dislocation motion to further enhance the extrinsic hardness through forming numerous atom-scale interlocks between different slip systems. Moreover, the HEDC maintains the excellent electrical conductivity of the constituent borides, and the competition between two correlating structures produces the special kind of coherent boundary that minimizes the scattering of conducting electrons and does not largely deteriorate the electrical conductivity.

Nanosensor-Driven Detection of Neuron-Derived Exosomal Aß42 with Graphene Electrolyte-Gated Transistor for Alzheimer's Disease Diagnosis.

Blood-based tests have sparked tremendous attention in non-invasive early diagnosis of Alzheimer's disease (AD), a most prevalent neurodegenerative malady worldwide. Despite significant progress in the methodologies for detecting AD core biomarkers such as Aß42 from serum/plasma, there remains cautious optimism going forward due to its controversial diagnostic value and disease relevance. Here, a graphene electrolyte-gated transistor biosensor is reported for the detection of serum neuron-derived exosomal Aß42 (NDE-Aß42), which is an emerging, compelling trove of blood biomarker for AD. Assisted by the antifouling strategy with the dual-blocking process, the noise against complex biological background was considerably reduced, forging an impressive sensitivity gain with a limit of detection of 447 ag/mL. An accurate detection of SH-SY5Y-derived exosomal Aß42 was also achieved with highly conformable enzyme-linked immunosorbent assay results. Importantly, the clinical analysis for 27 subjects revealed the immense diagnostic value of NDE-Aß42, which can outclass that of serum Aß42. The developed electronic assay demonstrates, for the first time, nanosensor-driven NDE-Aß42 detection, which enables a reliable discrimination of AD patients from non-AD individuals and even the differential diagnosis between AD and vascular dementia patients, with an accuracy of 100% and a Youden index of 1. This NDE-Aß42 biosensor defines a robust approach for blood-based confident AD ascertain.

Notch3 restricts metastasis of breast cancers through regulation of the JAK/STAT5A signaling pathway.

PURPOSE: To explore the potential role of signal transducer and activator of transcription 5A (STAT5A) in the metastasis of breast cancer, and its mechanism of regulation underlying. METHODS AND RESULTS: TCGA datasets were used to evaluate the expression of STAT5A in normal and different cancerous tissues through TIMER2.0, indicating that STAT5A level was decreased in breast cancer tissues compared with normal ones. Gene Set Enrichment Analysis predicted that STAT5A was associated with the activation of immune cells and cell cycle process. We further demonstrated that the infiltration of immune cells was positively associated with STAT5A level. Influorescence staining revealed the expression and distribution of F-actin was regulated by STAT5A, while colony formation assay, wound healing and transwell assays predicted the inhibitory role of STAT5A in the colony formation, migratory and invasive abilities in breast cancer cells. In addition, overexpression of the Notch3 intracellular domain (N3ICD), the active form of Notch3, resulted in the increased expression of STAT5A. Conversely, silencing of Notch3 expression by siNotch3 decreased STAT5A expression, supporting that STAT5A expression is positively associated with Notch3 in human breast cancer cell lines and breast cancer tissues. Mechanistically, chromatin immunoprecipitation showed that Notch3 was directly bound to the STAT5A promoter and induced the expression of STAT5A. Moreover, overexpressing STAT5A partially reversed the enhanced mobility of breast cancer cells following Notch3 silencing. Low expression of Notch3 and STAT5A predicted poorer prognosis of patients with breast cancer. CONCLUSION: The present study demonstrates that Notch3 inhibits metastasis in breast cancer through inducing transcriptionally STAT5A, which was associated with tumor-infiltrating immune cells, providing a novel strategy to treat breast cancer.

Rapid and precise detection of cancers via label-free SERS and deep learning.

Early, express, and reliable detection of cancer can provide a favorable prognosis and decrease mortality. Tumor biomarkers have been proven to be closely related to tumor occurrence and development. Conventional tumor biomarker detection based on genomic, proteomic, and metabolomic methods is time and equipment-consuming and always needs a specific target marker. Surface-enhanced Raman scattering (SERS), as a non-invasive ultrasensitive and label-free vibrational spectroscopy technique, can detect cancer-related biomedical changes in biofluids. In this paper, 110 serum samples were collected from 30 healthy controls and 80 cancer patients (including 30 bladder cancer (BC), 30 adrenal cancer (AC), and 20 acute myeloid leukemia (AML)). One microliter of blood serum was mixed with 1 µl silver colloid and then was air-dried for SERS measurements. After spectral data augmentation, one-dimensional convolutional neural network (1D-CNN) was proposed for precise and rapid identification of healthy and three different cancers with high accuracy of 98.27%. After gradient-weighted class activation mapping (Grad-CAM) based spectral interpretation, the contributions of SERS peaks corresponding to biochemical substances indicated the most potential biomarkers, i.e., L-tyrosine in bladder cancer; acetoacetate and riboflavin in adrenal cancer and phospholipids, amide-I, and α-Helix in acute myeloid leukemia, which might provide an insight into the mechanism of intelligent diagnosis of different cancers based on label-free serum SERS. The integration of label-free SERS and deep learning has great potential for the rapid, reliable, and non-invasive detection of cancers, which may significantly improve the precise diagnosis in clinical practice.

Acupuncture Needle-Based Transistor Neuroprobe for In Vivo Monitoring of Neurotransmitter.

Chemical communication via neurotransmitters is central to brain functions. Nevertheless, in vivo real-time monitoring of neurotransmitters released in the brain, especially the electrochemically inactive molecules, remains a great challenge. In this work, a novel needle field-effect transistor (FET) microsensor based on an acupuncture needle is proposed, which is demonstrated to be capable of real-time monitoring dopamine molecules as well as neuropeptide Y in vivo. The FET microstructure is fabricated by successively wrapping an insulating layer and a gold layer on the top of the needle, where the needle and the Au served as the source and drain, respectively. After assembling reduced graphene oxide (RGO) between the source and drain electrodes, the specific aptamer is immobilized on the RGO, making this needle-FET biosensor highly selective and sensitive to real-time monitor neurotransmitters released from rat brain, even in a Parkinson's diseases model. Furthermore, the needle-FET biosensor is applied to detect a variety of targets including hormones, proteins, and nucleic acid. By constructing a FET sensing interface on an acupuncture needle and implanting the sensor in a rat's brain for in vivo detection, this work provides a new sight in the FET domain and further expands the species of real-time in vivo detection.

RGD-functionalised melanin nanoparticles for intraoperative photoacoustic imaging-guided breast cancer surgery.

PURPOSE: Obtaining tumour-free margins is critical for avoiding re-excision and reducing local recurrence following breast-conserving surgery; however, it remains challenging. Imaging-guided surgery provides precise detection of residual lesions and assists surgical resection. Herein, we described water-soluble melanin nanoparticles (MNPs) conjugated with cyclic Arg-Gly-Asp (cRGD) peptides for breast cancer photoacoustic imaging (PAI) and surgical navigation. METHODS: The cRGD-MNPs were synthesised and characterized for morphology, photoacoustic characteristics and stability. Tumour targeting and toxicity of cRGD-MNPs were determined by using either breast cancer cells, MDA-MB-231 tumour-bearing mice or the FVB/N-Tg (MMTV-PyVT) 634Mul/J mice model. PAI was used to locate the tumour and guide surgical resection in MDA-MB-231 tumour-bearing mice. RESULTS: The cRGD-MNPs exhibited excellent in vitro and in vivo tumour targeting with low toxicity. Intravenous administration of cRGD-MNPs to MDA-MB-231 tumour-bearing mice showed an approximately 2.1-fold enhancement in photoacoustic (PA) intensity at 2 h, and the ratio of the PA intensity at the tumour site to that in the surrounding normal tissue was 3.2 ± 0.1, which was higher than that using MNPs (1.7 ± 0.3). Similarly, the PA signal in the spontaneous breast cancer increased ~ 2.5-fold at 2 h post-injection of cRGD-MNPs in MMTV-PyVT transgenic mice. Preoperative PAI assessed tumour volume and offered three-dimensional (3D) reconstruction images for accurate surgical planning. Surgical resection following real-time PAI showed high consistency with histopathological analysis. CONCLUSION: These results highlight that cRGD-MNP-mediated PAI provide a powerful tool for breast cancer imaging and precise tumour resection. cRGD-MNPs with fine PA properties have great potential for clinical translation.

Fabrication of tumor targeting rare-earth nanocrystals for real-time NIR-IIb fluorescence imaging-guided breast cancer precise surgery.

The near-infrared fluorescence imaging has been integrated into the operating room to guide tumor resection, potentially reducing the positive margin rates in breast-conserving surgery (BCS). Relative to the widely used first near-infrared fluorescence imaging, imaging in the second near-infrared (NIR-II) region possesses higher contrast and deeper tissue penetration, particularly in the NIR-IIb window, offering many new opportunities for imaging-guided BCS. Here, we fabricated the c(RGDfC) functionalized erbium-based rare-earth nanoparticles (ErNPs@cRGD) with superior optical property in NIR-IIb region. Owing to deeper tissue penetration and efficient tumor targeting, ErNPs@cRGD-based NIR-IIb fluorescence imaging achieved enhanced signal-to-background ratios in tumor visualization, which was able to guide more complete tumor resection, identify multiple microtumors and distinguish malignant lesions from normal tissues in various mice models. Based on these, this NIR-IIb imaging strategy for surgical navigation can significantly reduce positive margin rates and improve prognosis, laying a foundation for the clinical resection of breast cancer.

Ultrasensitive Exosomal MicroRNA Detection with a Supercharged DNA Framework Nanolabel.

DNA self-assembly has created various nanostructured probes deployed in biosensors, whereas their direct charge contribution to sensitive bioassays remains elusive. Here, we report a supercharged tetrahedral DNA nanolabel-based electrochemical (eTDN) sensor for ultrasensitive detection of exosomal microRNAs (Exo-miRs). By using an "assembly before testing" strategy, there is high-efficiency recognition between the target Exo-miR and self-assembled TDN probe in a homogenous solution relative to surface-based hybridization. The TDN-miR complex can be further bridged specifically to form a stable sandwich construct with the surface-confined capture sequence via the base-stacking effect. The intrinsic supercharges of the TDN can adsorb numerous electroactive molecules in stoichiometry, which largely enhances the detection sensitivity, particularly by using electroneutral peptide nucleic acid instead of DNA probes to minimize the background signal. Using this approach, the eTDN sensor achieves a high sensitivity (34 aM), high specificity (against the single mismatch), and high selectivity (in serum). Furthermore, this ultrasensitive sensor provides a conjugation-free, non-enzymatic Exo-miR detection in blood and accurately distinguishes the breast cancer patients from normal individuals, showing to be a promising tool in the early diagnosis of malignant tumors.

Carbon Nanotube Field-Effect Transistor Biosensor for Ultrasensitive and Label-Free Detection of Breast Cancer Exosomal miRNA21.

Tumor-derived exosomal miRNAs may have important functions in the onset and progression of cancers and are potential biomarkers for early diagnosis and prognosis monitoring. Yet, simple, sensitive, and label-free detection of exosomal miRNAs remains challenging. Herein, an ultrasensitive, label-free, and stable field-effect transistor (FET) biosensor based on a polymer-sorted high-purity semiconducting carbon nanotube (CNT) film is reported to detect exosomal miRNA. Different from conventional CNT FETs, the CNT FET biosensors employed a floating gate structure using an ultrathin Y2O3 as an insulating layer, and assembled Au nanoparticles (AuNPs) on Y2O3 as linkers to anchor probe molecules. A thiolated oligonucleotide probe was immobilized on the AuNP surface of the sensing area, after which miRNA21 was detectable by monitoring the current change before and after hybridization between the immobilized DNA probe and target miRNA. This method achieved both high sensitivity (LOD: 0.87 aM) and high specificity. Furthermore, the FET biosensor was employed to test clinical plasma samples, showing significant differences between healthy people and breast cancer patients. The CNT FET biosensor shows the potential applications in the clinical diagnosis of breast cancer.

Peptide Nucleic Acid-Functionalized Nanochannel Biosensor for the Highly Sensitive Detection of Tumor Exosomal MicroRNA.

Compared with free miRNAs in blood, miRNAs in exosomes have higher abundance and stability. Therefore, miRNAs in exosomes can be regarded as an ideal tumor marker for early cancer diagnosis. Here, a peptide nucleic acid (PNA)-functionalized nanochannel biosensor for the ultrasensitive and specific detection of tumor exosomal miRNAs is proposed. After PNA was covalently bound to the inner surface of the nanochannels, the detection of tumor exosomal miRNAs was achieved by the charge changes on the surface of nanochannels before and after hybridization (PNA-miRNA). Due to the neutral characteristics of PNA, the efficiency of PNA-miRNA hybridization was improved by significantly reducing the background signal. This biosensor could not only specifically distinguish target miRNA-10b from single-base mismatched miRNA but also achieve a detection limit as low as 75 aM. Moreover, the biosensor was further used to detect exosomal miRNA-10b derived from pancreatic cancer cells and normal pancreatic cells. The results indicate that this biosensor could effectively distinguish pancreatic cancer tumor-derived exosomes from the normal control group, and the detection results show good consistency with those of the quantitative reverse-transcription polymerase chain reaction method. In addition, the biosensor was used to detect exosomal miRNA-10b in clinical plasma samples, and it was found that the content of exosomal miRNA-10b in cancer patients was generally higher than that of healthy individuals, proving that the method is expected to be applied for the early diagnosis of cancer.

Iron, Nitrogen-Doped Carbon Aerogels for Fluorescent and Electrochemical Dual-Mode Detection of Glucose.

Due to their effective catalytic activity and maximum atom utilization, single metal atoms dispersed in carbon matrices have found diverse applications in electrocatalysis, photocatalysis, organic catalysis, and biosensing. Herein, iron is atomically dispersed into nitrogen-doped porous carbon aerogel by a facile pyrolysis procedure, and the resulting nanocomposite behaves both as a peroxidase mimic for the sensitive detection of glucose by fluorescence spectroscopy and as an effective catalyst for the electrochemical oxidation of glucose. The glucose concentration can be quantified within the millimolar to micromolar range with a limit of detection of 3.1 and 0.5 µM, respectively. Such a dual-functional detection platform also shows excellent reproducibility, stability, and selectivity, and the performance in glucose detection of clinical and artificial human body fluids is highly comparable to that of leading assays in recent studies and results from commercial sensors. Results from this study suggest that carbon aerogel-supported single atoms can be used as a dual-functional nanozyme for the construction of low-cost, high-performance dual-signal readout platforms for glucose detection.

Prevalence and Clinical Characteristics of Nontuberculous Mycobacteria in Patients with Bronchiectasis: A Systematic Review and Meta-Analysis.

BACKGROUND: Although international bronchiectasis guidelines recommended screening of nontuberculous mycobacteria (NTM) both at initial evaluation and prior to administration of macrolide treatment, data regarding NTM in bronchiectasis remain elusive. OBJECTIVE: To establish the prevalence, species, and clinical features of NTM in adults with bronchiectasis. METHODS: We searched PubMed, Embase, and Web of Science for studies published before April 2020 reporting the prevalence of NTM in adults with bronchiectasis. We only included studies with bronchiectasis confirmed by computed tomography and NTM identified by mycobacteria culture or molecular methods. Random-effects meta-analysis was employed. RESULTS: Of the 2,229 citations identified, 21 studies, including 12,454 bronchiectasis patients were included in the final meta-analysis. The overall pooled prevalence of NTM isolation and pulmonary NTM disease were 7.7% (5.0%-11.7%) (n/N = 2,677/12,454) and 4.1% (1.4%-11.4%) (n/N = 30/559), respectively, with significant heterogeneity (I2 = 97.7%, p < 0.001 and I2 = 79.9%, p = 0.007; respectively). The prevalence of NTM isolation varied significantly among different geographical regions with the highest isolation at 50.0% (47.3%-52.7%) reported in the United States. Mycobacterium avium complex and Mycobacterium abscessus complex accounted for 66 and 16.6% of all species, respectively. Some clinical and radiological differences were noted between patients with and without the presence of NTM isolation although the results are inconsistent. CONCLUSIONS: Heterogeneity in prevalence estimates of NTM isolation indicated that both local surveys to inform development of clinical services tailored to patients with bronchiectasis and population-based studies are needed. The clinical features associated with NTM in bronchiectasis and their incremental utility in studying the association is unknown and merits further investigation.

Inhibition of Notch1 reverses EMT and chemoresistance to cisplatin via direct downregulation of MCAM in triple-negative breast cancer cells.

Resistance to chemotherapy continues to be a critical issue in the clinical therapy of triple-negative breast cancer (TNBC). Epithelial-mesenchymal transition (EMT) is thought to contribute to chemoresistance in several cancer types, including breast cancer. Identification of the key signaling pathway that regulates the EMT program and contributes to chemoresistance in TNBC will provide a novel strategy to overcome chemoresistance in this subtype of cancer. Herein, we demonstrate that Notch1 positively associates with melanoma cell adhesion molecule (MCAM), a unique EMT activator, in TNBC tissue samples both at mRNA and protein levels. High expression of Notch1 and MCAM both predicts a poor survival in basal-like/TNBC patients, particularly in those treated with chemotherapy. The expression of Notch1 and MCAM in MDA-MB-231 cells gradually increases in a time-dependent manner when exposing to low dose cisplatin. Moreover, the expressions of Notch1 and MCAM in cisplatin-resistant MDA-MB-231 cells are significantly higher than wild-type counterparts. Notch1 promotes EMT and chemoresistance, as well as invasion and proliferation of TNBC cells via direct activating MCAM promoter. Inhibition of Notch1 significantly downregulates MCAM expression, resulting in the reversion of EMT and chemoresistance to cisplatin in TNBC cells. Our study reveals the regulatory mechanism of the Notch1 pathway and MCAM in TNBC and suggesting that targeting the Notch1/MCAM axis, in conjunction with conventional chemotherapies, might be a potential avenue to enhance the therapeutic efficacy for patients with TNBC.

Dual-Aptamer Modified Graphene Field-Effect Transistor Nanosensor for Label-Free and Specific Detection of Hepatocellular Carcinoma-Derived Microvesicles.

Cancerous microvesicles (MVs), which are heterogeneous membrane-bound nanovesicles shed from the surfaces of cancer cells into the extracellular environment, have been widely recognized as promising "biofingerprints" for various cancers. High-performance identification of cancerous MVs plays a vital role in the early diagnosis of cancer, yet it is still technically challenging. Herein, we report a gold nanoparticle (AuNP)-decorated, dual-aptamer modified reduced graphene oxide (RGO) field-effect transistor (AAP-GFET) nanosensor for the label-free, specific, and sensitive quantification of HepG2 cell-derived MVs (HepG2-MVs). After GFET chips were fabricated, AuNPs were then decorated on the RGO surface. For specific capture and detection of HepG2-MVs, both sulfhydrylated HepG2 cell specific TLS11a aptamer (AptTLS11a) and epithelial cell adhesion molecule aptamer (AptEpCAM) were immobilized on the AuNP surface through an Au-S bond. This developed nanosensor delivered a broad linear dynamic range from 6 × 105 to 6 × 109 particles/mL and achieved a high sensitivity of 84 particles/µL for HepG2-MVs detection. Moreover, this AAP-GFET platform was able to distinguish HepG2-MVs from other liver cancer-related serum proteins (such as AFP and CEA) and MVs derived from human normal cells and other cancer cells of lung, pancreas, and prostate, suggesting its excellent method specificity. Compared with those modified with a single type of aptamer alone (AptTLS11a or AptEpCAM), such an AAP-GFET nanosensor showed greatly enhanced signals, suggesting that the dual-aptamer-based bio-nano interface was uniquely designed and could realize more sensitive quantification of HepG2-MVs. Using this platform to detect HepG2-MVs in clinical blood samples, we found that there were significant differences between healthy controls and hepatocellular carcinoma (HCC) patients, indicating its great potential in early HCC diagnosis.

Multivalence-Actuated DNA Nanomachines Enable Bicolor Exosomal Phenotyping and PD-L1-Guided Therapy Monitoring.

Exosome-associated liquid biopsies are hampered by challenges in the exosomal quantification and phenotyping. Here, we present a bioinspired exosome-activated DNA molecular machine (ExoADM) with multivalent cyclic amplification that enables highly sensitive detection and phenotyping of circulating exosomes. ExoADM harbors two (an exposed and a hidden) DNA toehold domains that actuate sequential branch migration and multivalent recycling in response to exosomal surface markers. Importantly, this self-powered ExoADM achieves a high sensitivity (33 particles/µL) and is compatible with another DNA nanomachine targeting different exosomal surface markers for dual-color phenotyping. Using this strategy, we can simultaneously track the dynamic changes of ExoPD-L1 and ExoCD63 expression induced by signaling molecules. Further, we found that their expression levels on circulating exosomes could well differentiate cancer patients from the normal individuals. More importantly, ExoPD-L1 levels could reflect the efficacy of different treatments and guide anti-PD-1 immunotherapy, suggesting the potential of ExoPD-L1 in clinical diagnosis and targeted therapy monitoring.

Hierarchical Nanostructuring Array Enhances Mid-Hybridization for Accurate Herbal Identification via ITS2 DNA Barcode.

It remains a technical challenge to accurately identify close species of herbal medicines, especially from adulterants, because of their highly identical phenotypes and chemical compositions. Here, we report a direct, sequencing-free, high-curvature nanostructuring-based electrochemical herb sensor (nanoE-herb sensor) to identify herbal species quickly and accurately using ITS2 barcodes. We engineer a nano-roughened carbon-supported gold nanostructuring array by photolithograph-free, one-step electrodeposition. The 3D fractal nanostructures exhibit a high deflection angle that largely enhances DNA hybridization efficiency, particularly for the midcomplementary hybridization, as compared to the 2D planar surface. More importantly, such a trans-scale array biointerface (including macroscale carbon and nanoscale gold branches) can overcome the detection barrier of slow diffusion of a long genomic sequence and inaccessibility of the sequestered variations in ITS2 secondary structures through the out-protruded 3D functional nanostructures. Our nanoE-herb sensor achieves a detection limit of 0.18 fM for the 64-mer fragment of saffron ITS2 barcode with midhybridization and shows superior specificity against even single-base mismatch. The sensor also precisely differentiates saffron from six other adulterants by directly detecting unpurified asymmetric PCR amplicons (∼500 bp) with ITS2 sequences, suggesting its great potential in the field identification of herbal medicinal species and pathogenic bacteria with specific DNA barcodes.

Factors associated with prolonged viral shedding and impact of lopinavir/ritonavir treatment in hospitalised non-critically ill patients with SARS-CoV-2 infection.

BACKGROUND: The duration of viral shedding is central to the guidance of decisions about isolation precautions and antiviral treatment. However, studies regarding the risk factors associated with prolonged shedding of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the impact of lopinavir/ritonavir (LPV/r) treatment on viral shedding remain scarce. METHODS: Data were collected from all SARS-CoV-2 infected patients who were admitted to isolation wards and had reverse transcription PCR conversion at the No. 3 People's Hospital of Hubei province, China, between 31 January and 9 March 2020. We compared clinical characteristics and SARS-CoV-2 RNA shedding between patients initiated with LPV/r treatment and those without. Logistic regression analysis was employed to evaluate the risk factors associated with prolonged viral shedding. RESULTS: Of 120 patients, the median age was 52 years, 54 (45%) were male and 78 (65%) received LPV/r treatment. The median duration of SARS-CoV-2 RNA detection from symptom onset was 23 days (interquartile range 18-32 days). Older age (OR 1.03, 95% CI 1.00-1.05; p=0.03) and the lack of LPV/r treatment (OR 2.42, 95% CI 1.10-5.36; p=0.029) were independent risk factors for prolonged SARS-CoV-2 RNA shedding. Patients who initiated LPV/r treatment within 10 days from symptom onset, but not initiated from day 11 onwards, had significantly shorter viral shedding duration compared with those without LPV/r treatment (median 19 days versus 28.5 days; log-rank p<0.001). CONCLUSION: Older age and the lack of LPV/r treatment were independently associated with prolonged SARS-CoV-2 RNA shedding in patients with coronavirus disease 2019 (COVID-19). Earlier administration of LPV/r treatment could shorten viral shedding duration.

First Report of Pestalotiopsis lushanensis Causing Brown Leaf Spot on Sarcandra glabra in China.

Sarcandra glabra is a species of Chloranthaceae family and this family grow in the southern part of China, Japan, and Southeastern Asia (Li et al. 2019). It is a kind of precious Chinese herbal medicine, which occupies an important position in traditional Chinese herbal medicine. It plays an effective role in the treatment of cancer, rheumatism, pneumonia, digestive tract inflammations, traumatic injuries and fractures, anti-virus, anti-bacterial, antioxidant, etc. (Li et al. 2019; Zheng et al. 2003; Zhou et al. 2013). Since June 2020, we discovered a serious leaf disease in the S. glabra planting base of Shibing County (108.12E 27.03N), in Guizhou Province, with an incidence rate of 60% and yield losses of 40%. Initially, the symptoms developed as small specks where spots were purple with a dark brown halo margin, and round or oval. In later stages, the spots gradually expanded and became dry, whole severe leaf loss. To identify the pathogen, we collected the diseased leaves from S. glabra fields in Shibing County. Small tissue pieces from the edges of lesions were disinfected in 75% ethyl alcohol for 30 s and 1% hypochlorite for 1 min, rinsed five times in sterile water, plated on potato dextrose agar (PDA), and incubated at 28°C in lighted incubator for 3 days. Fungal colonies were consistently isolated and transferred to PDA for morphological characterization (Fang et al. 2007). Pathogenicity tests of the novel isolate HGUP CSH-2 were conducted by spraying spore suspensions with a concentration of 1.6×108 conidia/ml on surface-disinfected (70% ethyl alcohol, 30 s) leaves, while sterile distilled water was used as the control. Plants with inoculated leaves (three per treatment) were placed in lighted growth chambers at 28°C for 5 days and watered as needed (Light to dark ratio 1:1, RH=90%). Symptoms on inoculated leaves were similar to those described previously in the field. The same pathogenic fungus was re-isolated from the infected leaves but not from the non-inoculated leaves. Colonies on PDA attaining 70 mm diam after 7 d at 28°C, with pale honey-colored, sparse aerial mycelia on the surface with black, gregarious conidiomata. Conidiogenous cell discrete or integrated, ampulliform, clavate or subcylindrical, hyaline, smooth-walled, wide at base. Conidia fusoid, ellipsoid, straight to slightly curved, 4-septate, slightly constricted at septa, 22.26-27.17×6.9-8.22 µm (av.±SD: 24.68±1.57×7.68±0.38 µm; n=30). According to the colony and conidia characteristics, the isolate was initially identified as Pestalotiopsis spp. (Liu et al. 2017). The pathogen was confirmed by amplification and sequencing of the internal transcribed spacer region (ITS) gene, the translation elongation factor-1 (TEF1) gene and the ß-tubulin (TUB2) gene (Liu et al. 2017) using ITS1/ITS4, Bt2a/T1 and EF1-526F/1567R primers, respectively. The sequences of the PCR products were deposited in GenBank with accession numbers MT919215 (ITS), MT939300 (TUB2) and MT939299 (TEF1). BLAST results of the obtained sequences of the ITS, TUB2 and TEF1 genes revealed 97.16% (479/493 nucleotides), 99.56% (675/678 nucleotides) and 99.89% (890/891 nucleotides) homology with those of Pestalotiopsis lushanensis in GenBank (MG726538, KY464157 and KX895223). Maximum Likelihood method was used for phylogenetic analysis. The result showed that HGUP CSH-2 was together with P. lushanensis with a support rate of 100%. According to the morphological characteristics and molecular phylogenetic analysis, the pathogen was identified as P. lushanensis. So far as we know, our research is the first report of brown leaf spot of S. glabra caused by P. lushanensis in China. Thus, identification of P. lushanensis for this disease is important for the advancement of effective prevention and control practises as future perspectives.