Spatial-temporal evaluation of urban resilience in the Yangtze River Delta from the perspective of the coupling coordination degree.

Scientific evaluation of urban resilience will help to improve the ability of self-prevention and self-recovery when facing internal and external pressure. However, existing studies are on basis of the overall perspective of the urban resilience evaluation index system to measure urban resilience, often ignoring the coupling and coordination degree among indicators. Therefore, an empirical analysis is developed, which is used to measure the urban resilience of eight cities in the Yangtze River Delta urban agglomeration from 2010 to 2019 from the perspective of coupling coordination degree based on the urban resilience evaluation index system. The empirical results show that (1) In time, the eight cities' resilience fluctuated dynamically and varied to different degrees. It presents the spatial distribution characteristics of "high in the center and low in the periphery" in space. (2) In time, the coupling coordination degree in the eight cities fluctuated slightly. The spatial distribution pattern of "high in the center and low in the periphery" was formed in terms of space. (3) There is a long-term stable relationship between urban resilience and the coupling coordination degree among all indicators. In a certain sense, the higher the coupling coordination degree is, the higher the urban resilience is. These results can improve urban resilience to some extent and make cities more resilient in the future collaborative development process, and provide a way to evaluate urban resilience at different spatial-temporal scales.

A deep red ratiometric fluorescent probe for accurate detection of peroxynitrite in mitochondria.

Peroxynitrite (ONOO-) is widespread within living organisms and has been implicated in many physiological and pathological processes. Since ONOO- is mainly produced in mitochondria, accurate detection of ONOO- in mitochondria can help us understand its specific mechanism of action in the organism. Rather than single-wavelength emissive mitochondrial probes, ratiometric fluorescent probes with longer emission wavelength, large emission shift, and specific mitochondrial targeting properties are more likely to obtain a more accurate ONOO- content in mitochondria. To further avoid the interference by cytoplasmic ONOO-, we constructed a fluorescent probe MXMP with deep red emission and ratio properties, and it will be forbidden to enter the mitochondria after oxidation. In addition to its excellent selectivity and sensitivity, it shifted its fluorescence emission by up to 130 nm, with a detection limit of 84 nM.

Rhodol-derived turn-on fluorescent probe for copper ions with high selectivity and sensitivity.

A new rhodol-derived fluorescent probe 1 with picolinate as the recognition receptor was designed and simply synthesized using a one-step reaction. With the concentration of added Cu2+ increases, it gradually turns pink, so the effect of naked eye detection can be achieved. The detection limit of probe 1 for Cu2+ is 42 nM, and the linear detection range was 0-2 µM. The experimental results showed that 1 was a fluorescent probe with high selectivity, good water solubility, and high sensitivity to Cu2+ . Probe 1 was successfully applied in cell imaging experiments and can detect the concentration of Cu2+ in water samples. All these indicate that probe 1 has the potential to be applied to the detection of Cu2+ concentration in the real environment.

A duplex probe-directed recombinase amplification assay for detection of single nucleotide polymorphisms on 8q24 associated with prostate cancer

Single nucleotide polymorphisms (SNPs) have important application value in the research of population genetics, hereditary diseases, tumors, and drug development. Conventional methods for detecting SNPs are typically based on PCR or DNA sequencing, which is time-consuming, costly, and requires complex instrumentation. In this study, we present a duplex probe-directed recombinase amplification (duplex-PDRA) assay that can perform real-time detection of two SNPs (rs6983267 and rs1447295) in four reactions in two tubes at 39°C within 30 min. The sensitivity of duplex-PDRA was 2×103-104 copies per reaction and no cross-reactivity was observed. A total of 382 clinical samples (179 prostate cancer patients and 203 controls) from northern China were collected and tested by duplex-PDRA assay and direct sequencing. The genotyping results were completely identical. In addition, the association analysis of two SNPs with prostate cancer risk and bone metastasis was conducted. We found that the TT genotype of rs6983267 (OR: 0.42; 95%CI: 0.23-0.78; P=0.005) decreased the risk of prostate cancer, while the CA genotype of rs1447295 (OR: 1.89; 95%CI: 1.20-2.96; P=0.005) increased the risk of prostate cancer. However, no association between the two SNPs (rs6983267 and rs1447295) and bone metastasis in prostate cancer was found in this study (P>0.05). In conclusion, the duplex-PDRA assay is an effective method for the simultaneous detection of two SNPs and shows great potential for widespread use in research and clinical settings.

A duplex probe-directed recombinase amplification assay for detection of single nucleotide polymorphisms on 8q24 associated with prostate cancer.

Single nucleotide polymorphisms (SNPs) have important application value in the research of population genetics, hereditary diseases, tumors, and drug development. Conventional methods for detecting SNPs are typically based on PCR or DNA sequencing, which is time-consuming, costly, and requires complex instrumentation. In this study, we present a duplex probe-directed recombinase amplification (duplex-PDRA) assay that can perform real-time detection of two SNPs (rs6983267 and rs1447295) in four reactions in two tubes at 39°C within 30 min. The sensitivity of duplex-PDRA was 2×103-104 copies per reaction and no cross-reactivity was observed. A total of 382 clinical samples (179 prostate cancer patients and 203 controls) from northern China were collected and tested by duplex-PDRA assay and direct sequencing. The genotyping results were completely identical. In addition, the association analysis of two SNPs with prostate cancer risk and bone metastasis was conducted. We found that the TT genotype of rs6983267 (OR: 0.42; 95%CI: 0.23-0.78; P=0.005) decreased the risk of prostate cancer, while the CA genotype of rs1447295 (OR: 1.89; 95%CI: 1.20-2.96; P=0.005) increased the risk of prostate cancer. However, no association between the two SNPs (rs6983267 and rs1447295) and bone metastasis in prostate cancer was found in this study (P>0.05). In conclusion, the duplex-PDRA assay is an effective method for the simultaneous detection of two SNPs and shows great potential for widespread use in research and clinical settings.

Internally controlled recombinase-aided amplification (IC-RAA) assays for the detection of human papillomavirus genotypes 16 and 18 using extracted DNA and samples treated with nucleic acid releasing agent.

Cervical cancer is primarily caused by persistent infection with high-risk human papillomavirus (HPV), and 70% of cases are associated with HPV16 and 18 infections. The objective of this study was to establish rapid, simple, and sensitive internally controlled recombinase-aided amplification (IC-RAA) assays for the detection of HPV16 and 18. The assays were performed at 39 ℃ and were completed within 30 min. A total of 277 clinical samples of exfoliated cervical cells were tested by IC-RAA assays and commercial HPV real-time fluorescent PCR kits using extracted DNA and samples treated with nucleic acid releasing agent. The analytical sensitivity of the IC-RAA assay was found to be 10 copies/µL for the detection of HPV16 and 18 when using recombinant plasmids as targets. The optimal concentration of the internal control (IC) plasmid and 18 was 1000 copies/µL for HPV16 and 100 copies/µL for HPV18. The clinical sensitivity of the IC-RAA assays for HPV16 using extracted DNA and samples treated with nucleic acid releasing agent was 98.73% and 97.47%, respectively, with kappa values of 0.977 (P < 0.01) and 0.955 (P < 0.01), respectively, and 100% The specificity in both cases. For HPV18, the sensitivity and specificity were 100%, and the kappa value was 1 for both samples (P < 0.01). The IC-RAA assay is a promising tool for the detection of HPV16 and HPV18, especially in resource-constrained settings.

A coumarin-based fluorescent probe for Hg2+ and its application in living cells and zebrafish.

Mercury (Hg) is a heavy metal with high toxicity and easy migration; it can be enriched through the food chain, and cause serious threats to the natural environment and human health. So, the development of a method that can be used to detect mercury ions (Hg2+ ) in the environment, in cells, and in organisms is very important. Here, a new 7-hydroxycoumarin-derived carbonothioate-based probe (CC-Hg) was designed and synthesized for detection of Hg2+ . After addition of Hg2+ , a large fluorescence enhancement was observed due to the formation of 7-hydroxyl, which reinforced the intramolecular charge transfer process. The CC-Hg probe had good water solubility and selectivity. Moreover, the probe was able to detect Hg2+ quantitatively over the concentration range 0-2 µM and with a detection limit of 7.9 nM. Importantly, we successfully applied the probe to detect Hg2+ in water samples, in living cells, and in zebrafish. The experimental results demonstrated its potential value in practical applications.

Field applicable detection of hepatitis B virus using internal controlled duplex recombinase-aided amplification assay and lateral flow dipstick assay.

Hepatitis B virus (HBV) is a widespread blood-borne pathogen associated with the complication of liver cirrhosis and hepatocellular carcinoma, particularly in south-east Asian and African countries where HBV is highly endemic and the budget and resources are limited. Therefore, simple, rapid, and portable field detection methods are crucial to efficiently control HBV infection. In this study, using heat-treated DNA, we developed two-field applicable detection assays for HBV based on recombinase-aided amplification (RAA). One was an internal controlled duplex RAA assay using a portable real-time fluorescence detection device, another was an instrument-free visual observation assay using lateral flow dipsticks. The entire experimental time was greatly shortened to less than 40 minutes at 39.0°C. The sensitivities, specificities, and clinical performance of both assays were evaluated. Compared with quantitative polymerase chain reaction assay as a reference, our results demonstrated that the two RAA-based assay obtained 97.18% and 95.77% of sensitivity, respectively, and the specificity was 100%, by testing a total of 157 serum samples with HBsAg positive. We conclude that the advantages of rapidity, simplicity, portability, and visualization of proposed two assays make them great potentials in point-of-care testing of HBV infection by untrained people in resource-limited situations.

A highly sensitive one-tube nested quantitative real-time PCR assay for specific detection of Bordetella pertussis using the LNA technique.

OBJECTIVES: Bordetella pertussis is a highly contagious respiratory agent and is the causative pathogen of pertussis, which primarily affects children. Current diagnostic techniques for this pathogen have a variety of limitations including a long culture time, low bacterial load, and lack of specificity. METHODS: This article reports the development of a one-tube nested quantitative real-time PCR assay using the locked nucleic acid (LNA) technique (LNA-OTN-q-PCR), targeting the BP485 gene and using a simple inexpensive extraction method. A total of 130 clinical samples from patients with clinically suspected pertussis, collected from the Children's Hospital of Hebei, China, were tested by LNA-OTN-q-PCR assay. RT-PCR and two-step semi-nested PCR assays were performed in parallel for comparison. RESULTS: Only strains of B. pertussis were identified as positive, whereas all of the remaining strains were appropriately identified as negative by the LNA-OTN-q-PCR assay. A single copy per reaction can be detected by the LNA-OTN-q-PCR assay. Additionally, the sensitivity of this method was 100 times that of the RT-PCR assay (100 copies per reaction). Sixty-three of the 130 clinical samples were detected positive by LNA-OTN-q-PCR assay; in contrast, RT-PCR was able to detect only 41 positive samples. Following this, all 63 samples were positively identified by two-step semi-nested PCR. Compared with the two-step semi-nested PCR assay, both the specificity and sensitivity of the LNA-OTN-q-PCR assay using purified DNA and crude extract were 100%. CONCLUSIONS: This assay was able to detect B. pertussis infection with high sensitivity and specificity. This test shows great potential as a promising technique to detect B. pertussis in both clinical laboratories and public health settings.

A rapid and sensitive recombinase aided amplification assay incorporating competitive internal control to detect Bordetella pertussis using the DNA obtained by boiling.

OBJECTIVES: Pertussis is a highly transmissible acute respiratory infection caused by the bacterial pathogen Bordetella pertussis. The purpose of this study was to develop a rapid, simple and sensitive diagnostic test for detecting this pathogen. METHODS: Here we present a recombinase aided amplification (RAA) assay incorporating competitive internal amplification control (IAC) to detect Bordetella pertussis using the DNA obtained by boiling. This assay was performed in a single closed tube at 39°C within 30min. A total of 115 clinical samples suspected of pertussis were collected and tested by the internally controlled RAA assay using both extracted DNA with the commercial kit and the DNA obtained by boiling. For comparison, the real-time PCR (RT-PCR) was also performed with DNA extraction in parallel. RESULTS: The sensitivity of the internally controlled RAA assay was 101 copies or 10CFU/ml per reaction in detecting plasmid DNA or B. pertussis strain. The optimum concentration of the IAC plasmid was determined to be 100 copies, and the introduction of IAC effectively reduced the occurrence of false negatives. Compared to the RT-PCR, RAA results with DNA extraction obtained 100% sensitivity and specificity, and the RAA results with heat-treated DNA showed 85.96% sensitivity and 100% specificity. CONCLUSION: With the advantages of 45min turn-around time and simple steps of DNA purification, this assay could become a useful diagnostic tool for Bordetella pertussis detection and is potentially suitable for point-of-care identification to guide prompt clinical treatment.

A Highly Selective and Ultrasensitive Fluorescent Probe for Monitoring Hg2+ and Its Applications in Real Water Samples.

Mercury ions as high toxic pollutants have received wide-spread attention because of their poisonousness, persistence and enrichment. To better understand the distribution of mercury species and supplement more detailed toxicological research, it is necessary to develop some methods for monitoring mercury ions with high sensitivity and selectivity. Therefore, a simple rhodol-based highly selective fluorescent probe, RH-Hg, has been developed for monitoring Hg2+ with thiocarbamate as the recognition receptor. The probe RH-Hg can quantificationally detect mercury ions in aqueous solution assisted by hydrogen peroxide (H2O2), and it can discriminate Hg2+ through "naked-eye" observation of the color changes from light orange to dark pink. Finally, the practical applications of the probe RH-Hg in the river water further demonstrated that it will be an effective and economical tool for monitoring the distribution of Hg2+ in the environment.

A long-wavelength ultrasensitive colorimetric fluorescent probe for carbon monoxide detection in living cells.

Exploring techniques for monitoring the intracellular signaling molecule carbon monoxide (CO) in biosystems is important to help understand its various cellular functions. Therefore, a simple long-wavelength colorimetric fluorescent probe LW-CO was designed for selectively and sensitively detecting intracellular CO in living systems. Probe LW-CO is ultrasensitive and can track CO levels in the range of 0-1 µM, with a detection limit of about 3.2 nM. Additionally, the obvious color changes of probe LW-CO with CO (yellow to pink) provide a convenient way for on-site detection of CO with the naked eye. Probe LW-CO was applied to track the exogenous levels of CO in RAW264.7 cells. Probe LW-CO proved to be an efficient method for investigating various cellular functions of CO.

A carbonothioate-based far-red fluorescent probe for the specific detection of mercury ions in living cells and zebrafish.

The detection of ionic mercury (Hg2+) is very important because it is a highly toxic environmental pollutant that could cause serious diseases and threaten human health. Herein, we designed a new carbonothioate-based far-red fluorescent probe, CBRB, with a seminaphthorhodafluor dye as the fluorophore for the detection of Hg2+. The CBRB probe by itself exhibited very weak fluorescence due to the enhanced photo-induced electron transfer (PET) effect and inhibited the intramolecular charge transfer (ICT) process caused by the carbonothioate moiety. Upon addition of Hg2+, a tremendous fluorescence enhancement was achieved, attributed to the removal of the carbonothioate group via a specific mercury-promoted desulfurization reaction. Moreover, the probe displayed a large Stokes shift (about 105 nm) and was used to quantitatively measure the concentration of Hg2+ for concentrations ranging from 0 to 1 µM (DL = 3.6 nM). In addition, CBRB in our experiments responded exclusively to Hg2+, even in the presence of high concentrations other ions. Gratifyingly, this probe was successfully used to monitor Hg2+ in environmental water samples and to image Hg2+ in living cells as well as in zebrafish.

A highly selective and ultrasensitive ratiometric fluorescent probe for peroxynitrite and its two-photon bioimaging applications.

In this work, taking full advantage of the intramolecular charge transfer (ICT) mechanism, a hydroxynaphthalimide-based ratiometric two-photon fluorescent probe RTP-PN was synthesized to detect ONOO-. Probe RTP-PN could accurately detect ONOO- in the range of 1.4 nM-1.4 µM with the detection limit of 1.4 nM by a ratiometric fluorescence spectroscopy method. Additionally, probe RTP-PN exhibited an ultrafast response for ONOO- than other various species including H2O2 and ClO-. Finally, probe RTP-PN was successfully adopted to detect intracellular ONOO- by the two-photon excitation microscopy.

A highly specific and ultrasensitive p-aminophenylether-based fluorescent probe for imaging native HOCl in live cells and zebrafish.

It is very important to detect native hypochlorous acid (HOCl) in the complex biosystems owing to the important roles of HOCl in the immune defense and the pathogenesis of numerous diseases. In this paper, a new p-aminophenylether-based fluorescent probe PAPE-HA was developed for specific detection of HOCl. Probe PAPE-HA could implement the quantitative detection of HOCl ranging from 0 to 1 µM and the detection limit was obtained as low as 1.37 nM. Additionally, probe PAPE-HA could reach a rapid response for HOCl (<2 min). Importantly, probe PAPE-HA with preeminent specificity and ultrasensitivity was proven to possess powerful capability of tracking native HOCl in live cells and zebrafish, and we thus anticipate that probe PAPE-HA could be used as a novel promising tool for revealing diverse cellular functions of HOCl.

Applicability of duplex real time and lateral flow strip reverse-transcription recombinase aided amplification assays for the detection of Enterovirus 71 and Coxsackievirus A16.

BACKGROUND: Enterovirus 71 (EV71) and coxsackievirus A16 (CA16) are the two main etiological agents of Hand, Foot and Mouth Disease (HFMD). Simple and rapid detection of EV71 and CA16 is critical in resource-limited settings. METHODS: Duplex real time reverse-transcription recombinase aided amplification (RT-RAA) assays incorporating competitive internal amplification controls (IAC) and visible RT-RAA assays combined with lateral flow strip (LFS) for detection of EV71 and CA16 were developed respectively. Duplex real time RT-RAA assays were performed at 42 °C within 30 min using a portable real-time fluorescence detector, while LFS RT-RAA assays were performed at 42 °C within 30 min in an incubator. Recombinant plasmids containing conserved VP1 genes were used to analyze the sensitivities of these two methods. A total of 445 clinical specimens from patients who were suspected of being infected with HFMD were used to evaluate the performance of the assays. RESULTS: The limit of detection (LoD) of the duplex real time RT-RAA for EV71 and CA16 was 47 copies and 38 copies per reaction, respectively. The LoD of the LFS RT-RAA for EV71 and CA16 were both 91 copies per reaction. There was no cross reactivity with other enteroviruses. Compared to reverse transcription-quantitative PCR (RT-qPCR), the clinical diagnostic sensitivities of the duplex real time RT-RAA assay were 92.3% for EV71 and 99.0% for CA16, and the clinical diagnostic specificities were 99.7 and 100%, respectively. The clinical diagnostic sensitivities of the LFS RT-RAA assay were 90.1% for EV71 and 94.9% for CA16, and the clinical diagnostic specificities were 99.7 and 100%, respectively. CONCLUSIONS: The developed duplex real time RT-RAA and LFS RT-RAA assays for detection of EV71 and CA16 are potentially suitable in primary clinical settings.

Rational Design of a Hepatoma-Specific Fluorescent Probe for HOCl and Its Bioimaging Applications in Living HepG2 Cells.

Liver cancer is a kind of high mortality cancer due to the difficulty of early diagnosis. And according to the reports, the concentration of reactive oxygen species (ROS) was higher in cancer cells than normal cells. Therefore, developing an effective fluorescent probe for hepatoma-selective imaging of hypochlorous acid (HOCl) which is one of the vital ROS is of great importance for understanding the role of HOCl in liver cancer pathogenesis. However, the cell-selective fluorescent probe still remains a difficult task among current reports. Herein, a galactose-appended naphthalimide (Gal-NPA) with p-aminophenylether as a new receptor and galactose moiety as hepatoma targeting unit was synthesized and employed to detect endogenous HOCl in living HepG2 cells. This probe was proved to possess good water solubility and could respond specifically to HOCl. In addition, probe Gal-NPA could completely react to HOCl within 3 s meanwhile accompanied by tremendous fluorescence enhancement. The quantitative linear range between fluorescence intensities and the HOCl concentrations was 0 to 1 µM (detection limit = 0.46 nM). More importantly, fluorescence confocal imaging experiments showed that probe Gal-NPA could discriminate hepatoma cells over other cancer cells and simultaneously trace endogenous HOCl levels in living HepG2 cells. And we thus anticipate that probe Gal-NPA has the potential application for revealing the functions of HOCl in hepatoma cells.

A highly specific and ultrasensitive near-infrared fluorescent probe for imaging basal hypochlorite in the mitochondria of living cells.

The development of highly specific and ultrasensitive fluorescent probes for tracking basal mitochondrial hypochlorite is very important to unravel its diverse cellular functions in the mitochondria of living cells. In this paper, we have developed a water-soluble, mitochondria-targeted near-infrared fluorescent probe NB-OCl for selectively measuring OCl- in the presence of higher concentration (500 µM) other biologically important substances. Surprisingly, the obtained results demonstrated that probe NB-OCl could sensitively determine OCl- in the range of 0-200 pM with the detection limit of 10.8 pM. To the best of our knowledge, NB-OCl is the first fluorescent probe for the specific determination of OCl- at the picomolar level. Moreover, probe NB-OCl exhibits a fast response for OCl- (< 5 s), which would be in favor of tracking the highly reactive and short-lived OCl- in the living systems. The preeminent recognition properties of probe NB-OCl enable its applications in the monitoring of basal OCl- and the fluctuations of endogenous/exogenous OCl- levels in the mitochondria of living cells.

Rhodol-derived Colorimetric and Fluorescent Probe with the Receptor of Carbonothioate for the Specific Detection of Mercury Ions.

Developing some methods that can simply and effectively detect mercury ions (Hg2+) in the environment and biological systems are very important due to the problems of high toxicity and biological accumulation. Herein, we report a simple rhodol-derived colorimetric and fluorescent probe rhodol-Hg with a recognition receptor of carbonothioate for the specific determination of Hg2+. The color of probe rhodol-Hg solution changed remarkably from colorless to pink in the presence of Hg2+, thus rhodol-Hg could act as a "naked-eye" probe for Hg2+. Additionally, this probe exhibited high selectivity and ultrasensitivity in aqueous solution with the limit of detection (LOD) of 1.4 nM toward Hg2+, and the linear range was 0 - 0.8 µM determined by turn-on fluorescence spectrometry. Importantly, this probe has been successfully used for the detection of Hg2+ in environmental waters and living cells.

A highly specific and ultrasensitive fluorescent probe for basal lysosomal HOCl detection based on chlorination induced by chlorinium ions (Cl+).

The development of techniques for detecting HOCl at the subcellular level is very important to elucidate its cellular functions. Due to its relatively low concentration, it is still a great challenge to specifically track the basal HOCl in normal cells. In this paper, based on the unique chlorination of HOCl by the initiation of chlorinium ions (Cl+) in an acidic medium, we have developed a simple pH-mediated lysosome-targetable fluorescent probe Lyso-HOCl for the specific detection of HOCl over other bioactive molecules at higher concentration (500 µM). Our results show that Lyso-HOCl possesses a detection limit of 8.0 pM, and can quantitatively detect HOCl at the picomolar level. The ultrasensitive and ultrafast response property of probe Lyso-HOCl offers a good opportunity to monitor the basal HOCl and the fluctuation of endogenous HOCl levels in the lysosomes of macrophages (Raw 264.7 cells), and we thus anticipate that this probe would provide a promising tool for further unraveling the biological functions of HOCl in subcellular lysosomes.