The P2Y2 receptor mediates terminal adipocyte differentiation and insulin resistance: Evidence for a dual G-protein coupling mode.

Several P2Y nucleotide receptors have been shown to be involved in the early stage of adipocyte differentiation in vitro and insulin resistance in obese mice; however, the exact receptor subtype(s) and its underlying molecular mechanism in relevant human cells are unclear. Here, using human primary visceral preadipocytes as a model, we found that during preadipocyte-to-mature adipocyte differentiation, the P2Y2 nucleotide receptor (P2Y2R) was the most upregulated subtype among the eight known P2Y receptors and the only one further dramatically upregulated after inflammatory TNFα treatment. Functional studies indicated that the P2Y2R induced intracellular Ca2+, ERK1/2, and JNK signaling but not the p38 pathway. In addition, stimulation of the P2Y2R suppressed basal and insulin-induced phosphorylation of AKT, accompanied by decreased GLUT4 membrane translocation and glucose uptake in mature adipocytes, suggesting a role of P2Y2R in insulin resistance. Mechanistically, we found that activation of P2Y2R did not increase lipolysis but suppressed PIP3 generation. Interestingly, activation of P2Y2R triggered Gi-protein coupling, and pertussis toxin pretreatment largely inhibited P2Y2R-mediated ERK1/2 signaling and cAMP suppression. Further, treatment of the cells with AR-C 118925XX, a selective P2Y2R antagonist, significantly inhibited adipogenesis, and P2Y2R knockout decreased mouse body weight gain with smaller eWAT mass infiltrated with fewer macrophages as compared to WT mice in response to a Western diet. Thus, we revealed that terminal adipocyte differentiation and inflammation selectively upregulate P2Y2R expression and that P2Y2R mediates insulin resistance by suppressing the AKT signaling pathway, highlighting P2Y2R as a potential new drug target to combat obesity and type-2 diabetes.

The Yin and Yang of ERBB4: Tumor Suppressor and Oncoprotein.

ERBB4 (HER4) is a member of the ERBB family of receptor tyrosine kinases, a family that includes the epidermal growth factor receptor (EGFR/ERBB1/HER1), ERBB2 (Neu/HER2), and ERBB3 (HER3). EGFR and ERBB2 are oncoproteins and validated targets for therapeutic intervention in a variety of solid tumors. In contrast, the role that ERBB4 plays in human malignancies is ambiguous. Thus, here we review the literature regarding ERBB4 function in human malignancies. We review the mechanisms of ERBB4 signaling with an emphasis on mechanisms of signaling specificity. In the context of this signaling specificity, we discuss the hypothesis that ERBB4 appears to function as a tumor suppressor protein and as an oncoprotein. Next, we review the literature that describes the role of ERBB4 in tumors of the bladder, liver, prostate, brain, colon, stomach, lung, bone, ovary, thyroid, hematopoietic tissues, pancreas, breast, skin, head, and neck. Whenever possible, we discuss the possibility that ERBB4 mutants function as biomarkers in these tumors. Finally, we discuss the potential roles of ERBB4 mutants in the staging of human tumors and how ERBB4 function may dictate the treatment of human tumors. SIGNIFICANCE STATEMENT: This articles reviews ERBB4 function in the context of the mechanistic model that ERBB4 homodimers function as tumor suppressors, whereas ERBB4-EGFR or ERBB4-ERBB2 heterodimers act as oncogenes. Thus, this review serves as a mechanistic framework for clinicians and scientists to consider the role of ERBB4 and ERBB4 mutants in staging and treating human tumors.

Carba PBP: a novel penicillin-binding protein-based lateral flow assay for rapid phenotypic detection of carbapenemase-producing Enterobacterales.

Rapid phenotypic detection assays, including Carba NP and its variants, are widely applied for clinical diagnosis of carbapenemase-producing Enterobacterales (CPE). However, these tests are based on the acidification of the pH indicator during carbapenem hydrolysis, which limits test sensitivity and speed, especially for the detection of CPE producing low-activity carbapenem (e.g., OXA-48 variants). Herein, we developed a novel rapid and sensitive CPE detection method (Carba PBP) that could measure substrate (meropenem) consumption based on penicillin-binding protein (PBP). Meropenem-specific PBP was used to develop a competitive lateral flow assay (LFA) for meropenem identification. For the detection of carbapenemase activity, meropenem concentration was optimized using a checkerboard assay. The performance of Carba PBP was evaluated and compared with that of Carba NP using a panel of 94 clinical strains characterized by whole-genome sequencing and carbapenem susceptibility test. The limit of detection of PBP-based LFA for meropenem identification was 7 ng mL-1. Using 10 ng mL-1 meropenem as the substrate, Carba PBP and Carba NP could detect 10 ng mL-1 carbapenemase within 25 min and 1,280 ng mL-1 CPE in 2 h, respectively. The sensitivity and specificity were 100% (75/75) and 100% (19/19) for Carba PBP and 85.3% (64/75) and 100% (19/19) for Carba NP, respectively. When compared with Carba NP, Carba PBP showed superior performance in detecting all the tested CPE strains (including OXA-48-like variants) within 25 min and presented two orders of magnitude higher analytical sensitivity, demonstrating potential for clinical diagnosis of CPE. IMPORTANCE This study successfully achieved the goal of carbapenemase activity detection with both high sensitivity and convenience, offering a convenient lateral flow assay for clinical diagnosis of carbapenemase-producing Enterobacterales.

Optimal dietary zinc inclusion improved growth performance, serum antioxidant capacity, immune status, and liver lipid and glucose metabolism of largemouth bass (Micropterus salmoides).

This study was conducted to ascertain the effect of dietary Zn on growth and health status of juvenile largemouth bass (Micropterus salmoides). Six experimental diets with Zn level of 50.17, 56.74, 73.34, 86.03, 123.94, and 209.20 mg/kg, respectively were compounded using complex amino acid-chelated zinc, and were fed to juvenile fish (5.50 ± 0.10 g) for 70 d. The specific growth rate (SGR) varied with dietary Zn level in a quadratic model and peaked at the 73.34 mg/kg group, while the feeding rate exhibited an opposite trend (P < 0.05). The condition factor, hepatosomatic index and mesenteric fat index all exhibited a tendency similar with SGR (P < 0.05). Dietary Zn level affected serum total proteins, urea, triglycerides, and glucose (P < 0.05). Serum Zn and copper levels linearly increased with dietary Zn level, while serum iron and manganese showed the opposite trend. Serum superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) increased with dietary Zn level and reached a plateau at 86.03 mg/kg. Serum complement component 3 (C3), IgM, and lysozyme also were enhanced by 73.34 mg/kg Zn. Body protein content increased with zinc level up to 73.34 mg/kg, and then remained steadily. As dietary Zn level increased, hepatic lipid level increased and then reached a plateau at 86.03 mg/kg group, while glycogen increased linearly. Moreover, gene expression related to lipid and glycogen metabolism from liver transcriptome further explained the liver lipid and glycogen variations. To conclude, a dietary Zn requirement of 76.99 mg/kg was suggested for juvenile largemouth bass to improve growth, antioxidant capacity, and immune status.

A comprehensive review on the detection of Staphylococcus aureus enterotoxins in food samples.

Staphylococcal enterotoxins (SEs), the major virulence factors of Staphylococcus aureus, cause a wide range of food poisoning and seriously threaten human health by infiltrating the food supply chain at different phases of manufacture, processes, distribution, and market. The significant prevalence of Staphylococcus aureus calls for efficient, fast, and sensitive methods for the early detection of SEs. Here, we provide a comprehensive review of the hazards of SEs in contaminated food, the characteristic and worldwide regulations of SEs, and various detection methods for SEs with extensive comparison and discussion of benefits and drawbacks, mainly including biological detection, genetic detection, and mass spectrometry detection and biosensors. We highlight the biosensors for the screening purpose of SEs, which are classified according to different recognition elements such as antibodies, aptamers, molecularly imprinted polymers, T-cell receptors, and transducers such as optical, electrochemical, and piezoelectric biosensors. We analyzed challenges of biosensors for the monitoring of SEs and conclude the trends for the development of novel biosensors should pay attention to improve samples pretreatment efficiency, employ innovative nanomaterials, and develop portable instruments. This review provides new information and insightful commentary, important to the development and innovation of further detection methods for SEs in food samples.

A Proof-of-Concept Sandwich Enzyme-Linked Immunoassay Development for Small Molecules.

A sandwich immunoassay theoretically exhibits higher sensitivity and specificity compared to a competitive counterpart; however, it is extremely difficult to obtain a pair of antibodies that can bind to a small molecule simultaneously, which is always thought to be a single epitope. In the present study, abamectin (ABM) was selected to prove the effect of hapten design and antibody recognition properties on the development of a sandwich immunoassay for small molecules. First, the epitopes of ABM were roughly located, and epitope distances were determined. Then, two haptens were designed by introducing spacer arms at the C4″-OH and C5-OH of ABM, respectively, aiming to provide the longest epitope distances. A total of seven rabbit polyclonal antibodies (pAbs) and 21 mouse monoclonal antibodies (mAbs) with various recognition properties were obtained. Extensive combinatorial associations of antibody pairs for simultaneously binding to ABM were performed, and only two mAb-mAb pairs were observed to achieve a sandwich immunoassay for ABM with a total success rate of 0.27%. The best mAb pair for sandwich immunoassay was confirmed by surface plasmon resonance, used to develop a sandwich immunoassay, and then evaluated by cross-reactivities and molecular docking with structurally similar analogues and abamectin. Altogether, the study provided a theoretical foundation as well as practical experience and demonstrated the importance of careful hapten design and extensive antibody screening to successfully establish the sandwich immunoassay for small molecules.

How Exactly Do AIEgens Target Bacteria? Leveraging the Targeting Mechanism to Design Sensitive Fluorescent Immunosensors.

Aggregation-induced emission luminogens (AIEgens) are promising candidates for bacterial imaging and detection because they can "Light-Up" pathogenic bacteria without complicated labeling or washing steps. However, there have been few in-depth analyses of the intrinsic mechanism underlying their utility as fluorescence probes for targeting bacteria. Therefore, using large-scale molecular dynamics simulations, we investigated the mechanism of their bacterial "Light-Up" behavior with N,N-diphenyl-4-(7-(pyridin-4-yl)benzo[c][1,2,5]thiadiazol-4-yl) aniline functionalized with 1-bromoethane (TBP-1). We propose that the triphenylamine motif of TBP-1, rather than the positively charged pyridine group, first contacts the cell membrane. After TBP-1 completely inserts into the cell membrane, the hydrophobic triphenylamine motif localizes in the hydrophobic core of the cell membrane, restricting the molecular variation of TBP-1, which induces the fluorescent "turn-on" and bacterial "Light-Up." On this basis, we established a heterogeneous lateral flow immunoassay (LFIA) for the detection of foodborne pathogens. The LFIA system showed improved sensitivity with a limit of detection as low as 103 CFU mL-1 and strong specificity. Our protocol opened an effective shortcut to the design of more efficient AIEgens and novel AIEgens-based immunoassays.

Involvement of the inhibition of mitochondrial apoptotic, p53, NF-κB pathways and the activation of Nrf2/HO-1 pathway in the protective effects of curcumin against copper sulfate-induced nephrotoxicity in mice.

Chronic copper exposure could cause potential nephrotoxicity and effective therapy strategies are limited. This study investigated the protective effects of curcumin on copper sulfate (CuSO4)-induced renal damage in a mouse model and the underlying molecular mechanisms. Mice were administrated orally with CuSO4 (100 mg/kg per day) in combination with or without curcumin (50, 100 or 200 mg/kg per day, orally) for 28 days. Results showed that curcumin supplementation significantly reduce the Cu accumulation in the kidney tissues of mice and improved CuSO4-induced renal dysfunction. Furthermore, curcumin supplantation also significantly ameliorated Cu exposure-induced oxidative stress and tubular necrosis in the kidneys of mice. Moreover, compared to the CuSO4 alone group, curcumin supplementation at 200 mg/kg per day significantly decreased CuSO4-induced the expression of p53, Bax, IL-1ß, IL-6, and TNF-α proteins, levels of NF-κB mRNA, levels of caspases-9 and - 3 activities, and cell apoptosis, and significantly increased the levels of Nrf2 and HO-1 mRNAs in the kidney tissues. In conclusion, for the first time, our results reveal that curcumin could trigger the inhibition of oxidative stress, mitochondrial apoptotic, p53, and NF-κB pathways and the activation of Nrf2/HO-1 pathway to ameliorate Cu overload-induced nephrotoxicity in a mouse model. Our study highlights that curcumin supplementation may be a promising treatment strategy for treating copper overload-caused nephrotoxicity.

MC-PRPA-HLFIA Cascade Detection System for Point-of-Care Testing Pan-Drug-Resistant Genes in Urinary Tract Infection Samples.

Recently, urinary tract infection (UTI) triggered by bacteria carrying pan-drug-resistant genes, including carbapenem resistance gene blaNDM and blaKPC, colistin resistance gene mcr-1, and tet(X) for tigecycline resistance, have been reported, posing a serious challenge to the treatment of clinical UTI. Therefore, point-of-care (POC) detection of these genes in UTI samples without the need for pre-culturing is urgently needed. Based on PEG 200-enhanced recombinase polymerase amplification (RPA) and a refined Chelex-100 lysis method with HRP-catalyzed lateral flow immunoassay (LFIA), we developed an MCL-PRPA-HLFIA cascade assay system for detecting these genes in UTI samples. The refined Chelex-100 lysis method extracts target DNA from UTI samples in 20 min without high-speed centrifugation or pre-incubation of urine samples. Following optimization, the cascade detection system achieved an LOD of 102 CFU/mL with satisfactory specificity and could detect these genes in both simulated and actual UTI samples. It takes less than an hour to complete the process without the use of high-speed centrifuges or other specialized equipment, such as PCR amplifiers. The MCL-PRPA-HLFIA cascade assay system provides new ideas for the construction of rapid detection methods for pan-drug-resistant genes in clinical UTI samples and provides the necessary medication guidance for UTI treatment.

Mobile Oxazolidinone Resistance Genes in Gram-Positive and Gram-Negative Bacteria.

Seven mobile oxazolidinone resistance genes, including cfr, cfr(B), cfr(C), cfr(D), cfr(E), optrA, and poxtA, have been identified to date. The cfr genes code for 23S rRNA methylases, which confer a multiresistance phenotype that includes resistance to phenicols, lincosamides, oxazolidinones, pleuromutilins, and streptogramin A compounds. The optrA and poxtA genes code for ABC-F proteins that protect the bacterial ribosomes from the inhibitory effects of oxazolidinones. The optrA gene confers resistance to oxazolidinones and phenicols, while the poxtA gene confers elevated MICs or resistance to oxazolidinones, phenicols, and tetracycline. These oxazolidinone resistance genes are most frequently found on plasmids, but they are also located on transposons, integrative and conjugative elements (ICEs), genomic islands, and prophages. In these mobile genetic elements (MGEs), insertion sequences (IS) most often flanked the cfr, optrA, and poxtA genes and were able to generate translocatable units (TUs) that comprise the oxazolidinone resistance genes and occasionally also other genes. MGEs and TUs play an important role in the dissemination of oxazolidinone resistance genes across strain, species, and genus boundaries. Most frequently, these MGEs also harbor genes that mediate resistance not only to antimicrobial agents of other classes, but also to metals and biocides. Direct selection pressure by the use of antimicrobial agents to which the oxazolidinone resistance genes confer resistance, but also indirect selection pressure by the use of antimicrobial agents, metals, or biocides (the respective resistance genes against which are colocated on cfr-, optrA-, or poxtA-carrying MGEs) may play a role in the coselection and persistence of oxazolidinone resistance genes.

Minimum Distance Between Two Epitopes in Sandwich Immunoassays for Small Molecules.

The pursuit of the limit between dimensionalities is a scientific goal with high applicability. Sandwich immunoassay, usually based on two antibodies binding two epitopes, is one of the most popular mainstay tools in both academic and industrial fields. Herein, we determined and evaluated the minimum distance of two epitopes in sandwich immunoassays for small molecules. Briefly, nine model analytes comprising two hapten epitopes, that is, melamine (MEL) and p-nitroaniline (NIA), were designed by increasing the linear chain linkers brick by brick. Two groups of monoclonal antibodies (mAbs) were produced with different recognition properties toward MEL and NIA using 12 new haptens with different spacer arms. The results indicated that two epitopes of the analyte with a distance of only 2.4 Å could be simultaneously bound by two mAbs, which is the known limit of epitope distance in sandwich immunoassays thus far. We further found that an epitope distance of below 8.8 Å for the analyte generally induces noticeable steric hindrance of antibodies, preventing a sandwich immunoassay with high probability. These observations were investigated and evaluated by molecular docking, molecular dynamics, and surface plasmon resonance and using model and real analytes. Altogether, we determined the minimum distance of two epitopes and explored the molecular mechanism of the antibody-analyte-antibody ternary complex in sandwich immunoassays, providing a theoretical basis for hapten design, antibody discovery and development, and sandwich immunoassay establishment for small molecules.

Synergy of outer membrane disruptor SLAP-S25 with hydrophobic antibiotics against Gram-negative pathogens.

OBJECTIVES: An effective strategy for combating MDR Gram-negative pathogens can greatly reduce the cost and shorten the antibiotic development progress. Here, we investigated the synergistic activity of outer membrane disruptor SLAP-S25 in combination with hydrophobic antibiotics (LogP > 2, including novobiocin, erythromycin, clindamycin and rifampicin) against MDR Gram-negative pathogens. METHODS: Five representative Gram-negative bacteria were selected as model strains to analyse the synergistic combination of SLAP-S25 and hydrophobic antibiotics. Carbapenem-resistant hypervirulent Klebsiella pneumoniae CRHvKP4 was used to investigate the synergistic mechanism. The in vivo synergistically therapeutic activity of SLAP-S25 and hydrophobic antibiotics was measured in the mouse peritonitis/sepsis model infected with K. pneumoniae CRHvKP4. RESULTS: SLAP-S25 disrupted the outer membrane by removing LPS from Gram-negative bacteria, facilitating the entry of hydrophobic antibiotics to kill MDR Gram-negative pathogens. Moreover, the combination of SLAP-S25 and rifampicin exhibited promising therapeutic effects in the mouse infection model infected with K. pneumoniae CRHvKP4. CONCLUSIONS: Our findings provide a potential therapeutic strategy to combine SLAP-S25 with hydrophobic antibiotics for combating MDR Gram-negative pathogens.

A rare monoclonal antibody discovery based on indirect competitive screening of a single hapten-specific rabbit antibody secreting cell.

A rare antibody that is able to tolerate physio-chemical factors is preferred and highly demanded in diagnosis and therapy. Rabbit monoclonal antibodies (RmAbs) are distinguished owing to their high affinity and stability. However, the efficiency and availability of traditional methods for RmAb discovery are limited, particularly for small molecules. Here, we present an indirect competitive screening method in nanowells, named CSMN, for single rabbit antibody-secreting cells (ASCs) selection with 20.6 h and propose an efficient platform for RmAb production against small molecules within 5.8 days for the first time. Chloramphenicol (CAP) as an antibacterial agent poses a great threat to public health. We applied CSMN to select CAP-specific ASCs and produced one high-affinity RmAb, surprisingly showed extremely halophilic properties with an IC50 of 0.08 ng mL-1 in the saturated salt solution, which has not yet been seen for other antibodies. The molecular dynamic simulation showed that the negatively charged surface improved the stability of the RmAb structure with additional disulfide bonds compared with mouse antibodies. Moreover, the reduced solvent accessible surface area of the binding pocket increased the interactions of RmAb with CAP in a saturated salt solution. Furthermore, RmAb was used to develop an immunoassay for the detection of CAP in real biological samples with simple pretreatment, shorter assay time, and higher sensitivity. The results demonstrated that the practical and efficient CSMN is suitable for rare RmAb discovery against small molecules.

Equisetin Targets Intracellular Staphylococcus aureus through a Host Acting Strategy.

Mammalian cells act as reservoirs of internalized bacteria to circumvent extracellular antibacterial compounds, resulting in relapse and reinfection diseases. The intracellular persistence of Staphylococcus aureus renders most traditional antibiotics useless, due to their inadequate subcellular accumulation. To replenish our antibiotic arsenal, we found that a marine-derived compound, equisetin, efficiently eliminates intracellular S. aureus by potentiating the host autophagy and inducing mitochondrial-mediated ROS generation to clear the invading S. aureus. The remarkable anti-infection activity of equisetin was validated in a peritonitis-infected mouse model. The marine product equisetin utilizes a unique dual mechanism to modulate the host-pathogen interaction in the clearance of intracellular bacteria. Thus, equisetin is an inspiring host-acting candidate for overcoming intracellular pathogens.

Prevalence of mcr-1 in Colonized Inpatients, China, 2011-2019.

In response to the spread of colistin resistance gene mcr-1, China banned the use of colistin in livestock fodders. We used a time-series analysis of inpatient colonization data from 2011-2019 to accurately reveal the associated fluctuations of mcr-1 that occurred in inpatients in response to the ban.

Engineering of Organic Solvent-Tolerant Antibody to Sulfonamides by CDR Grafting for Analytical Purposes.

The use of organic solvents to extract chemical contaminants for an immunoassay is mostly inevitable. On this occasion, the intolerance of natural antibodies against organic solvent is detrimental to the performance of immunoassays in terms of sensitivity, assay time, accuracy, and precision. Few studies have focused on improving the low tolerance of natural antibodies to organic solvents for analytical purposes. In this study, we engineered the monoclonal antibody (mAb) 4D11 to sulfonamides through CDR grafting by using one proven highly stable humanized antibody (hAb) 4D5 for the first time. The engineered antibody hAb 4D11 showed significantly improved tolerance abilities to acetonitrile (2% to 20%) and methanol (10% to 20%), and retained the highly affinity and class-specificity to sulfonamides. This study provided a general strategy to improve antibody tolerance to organic solvents and was greatly beneficial to the robust development of immunoassays.

Portable Magnetofluidic Device for Point-of-Need Detection of African Swine Fever.

With a nearly 100% mortality rate, African swine fever (ASF) has devastated the pork industry in many countries. Without a vaccine in sight, mitigation rests on rapid diagnosis and immediately depopulating infected or exposed animals. Unfortunately, current tests require centralized laboratories with well-trained personnel, take days to report the results, and thus do not meet the need for such rapid diagnosis. In response, we developed a portable, sample-to-answer device that allows for ASF detection at the point of need in <30 min. The device employs droplet magnetofluidics to automate DNA purification from blood, tissue, or swab samples and utilizes fast thermal cycling to perform real-time quantitative polymerase chain reaction (qPCR), all within an inexpensive disposable cartridge. We evaluated its diagnostic performance at six farms and slaughter facilities. The device exhibits high diagnostic accuracy with a positive percent agreement of 92.2% and a negative percent agreement of 93.6% compared with a lab-based reference qPCR test.

Rapid detection of human origin colistin-resistance genes mcr-1, mcr-3, mcr-8, mcr-10 in clinical fecal samples.

Plasmid-mediated colistin-resistance genes have been reported in human origin clinical samples worldwide which raises its threats to human infections. Notably, mcr-1, mcr-3, mcr-8, and mcr-10 have been reported isolated directly from clinical samples which creates more seriously threaten to human health than other mcr gene types. A multiplex polymerase chain reaction (Multi-PCR) protocol was developed to detect and genotype mobile colistin-resistance genes (mcr-1, mcr-3, mcr-8, mcr-10) in Enterobacteria for clinical laboratory purposes. We first designed four pairs of new primers for the amplification of mcr-1, mcr-3, mcr-8, and mcr-10 gene respectively to achieve stepwise separation of amplicons between 216 and 241 bp, and complete this Multi-PCR system with the assistance of another pair of universal primer. Among which the forward primers for mcr-8 and mcr-10 amplicons were identical. The protocol was validated by testing 11 clinical isolates of Escherichia coli and 3 clinical isolates of Klebsiella from human origin, each well characterized and prospectively validated. The Multi-PCR assay showed full concordance with whole-genome sequence data and displayed higher sensitivity and 100% specificity. The assay could detect all variants of the various mcr alleles described. The Multi-PCR assay successfully genotyped of mcr alleles described in one test.

Production of highly sensitive monoclonal antibody and development of lateral flow assays for phallotoxin detection in urine.

Phallotoxins, toxic cyclopeptides found in wild poisonous mushrooms, are predominant causes of fatal food poisoning. For the early and rapid diagnosis mushroom toxin poisoning, a highly sensitive and robust monoclonal antibody (mAb) against phallotoxins was produced for the first time. The half-maximum inhibition concentration (IC50) values of the mAb-based indirect competitive ELISAs for phallacidin (PCD) and phalloidin (PHD) detection were 0.31 ng mL-1 and 0.35 ng mL-1, respectively. In response to the demand for rapid screening of the type of poisoning and accurate determination of the severity of poisoning, colloidal gold nanoparticle (GNP) and time-resolved fluorescent nanosphere (TRFN) based lateral flow assays (LFA) were developed. The GNP-LFA has a visual cut-off value of 3.0 ng mL-1 for phallotoxins in human urine sample. The TRFN-LFA provides a quantitative readout signal with detection limit of 0.1 ng mL-1 in human urine sample. In this study, urine samples without pretreatment were used directly for the LFA strip tests, and both two LFAs were able to accomplish analysis within 10 min. The results demonstrated that LFAs based on the newly produced, highly sensitive, and robust mAb were able to be used for both rapid qualitative screening of the type of poisoning and accurate quantitative determination of the severity of poisoning after accidental ingestion by patients of toxic mushrooms.

Estimating aquatic plant diversity and distribution in rivers from Jingjinji region, China, using environmental DNA metabarcoding and a traditional survey method.

Traditional survey methods (TSMs) are difficult to use to perform a census of aquatic plant diversity completely in river ecosystems, and improved aquatic plant community monitoring programs are becoming increasingly crucial with a continuous decline in diversity. Although environmental DNA (eDNA) metabarcoding has been applied successfully to assess aquatic biodiversity, limited work has been reported regarding aquatic plant diversity in rivers. In this study, the efficiency of eDNA to estimate the aquatic plant diversity and spatial distribution of rivers from the Jingjinji (JJJ) region was evaluated by comparing results obtained by the TSM. Based on a combination of the two methods, 157 aquatic plant species, including 24 hydrophytes, 61 amphibious plants, and 72 mesophytes, were identified. The spatial patterns in species richness and abundance by eDNA exhibited agreement with the TSM results with a gradual decline from the mountain area (MA) to the agricultural area (AA) and then to the urban area (UA). Compared to the TSM, eDNA identified a significantly greater number of species per site (p < 0.01) and obtained a significantly higher abundance in hydrophytes (p < 0.01), supplementing the unavailable abundance data from the TSM. Furthermore, the aquatic plant assemblages from the different areas were discriminated well using eDNA (p < 0.05), but they were better discriminated by the TSM (p < 0.01). Thus, our study provides more detailed data on aquatic plant diversity in rivers from the JJJ region, which is essential for biodiversity conservation. Our findings also highlight that eDNA can be reliable for evaluating aquatic plant diversity and has the potential to respond to landscape heterogeneity in river ecosystems.